Copyright 0 1991 by the Genetics Society of America
Fine Mapping of Satellite DNA Sequences Along theY Chromosome of
Drosophila melanogastec Relationships Between Satellite Sequences and
Fertility Factors
Silvia Bonaccorsi* and AllanLohet
*Centro di Genetica Evolurionistica del CNR, Universita di Roma “La Sapienza,” 00185 Roma, Italy, and ?Department of Genetics,
Washington University School of Medicine, St. Louis, Missouri 63130
Manuscript received February 11, 1991
Accepted for publication May 29, 1991
ABSTRACT
The entirely heterochromatic Y chromosome of Drosophila melanogaster contains a series of simple
sequence satellite DNAs which together account for about 80% of its length. Molecular cloning of
the three simple sequence satellite DNAs of D. melanogaster (1.672, 1.686 and 1.705 g/ml) revealed
that each satellite comprises several distinct repeat sequences. Together 11 related sequences were
identified and 9 of them were shown to be located on the Y chromosome. In the present study we
have finely mapped 8 of these sequences along the Y by in situ hybridization on mitotic chromosome
preparations. The hybridization experiments were performed on a series of cytologicallydetermined
rearrangements involving the Y chromosome. The breakpoints of these rearrangements provided an
array of landmarks along the Y whichhave been used to localize each sequence on the various
heterochromatic blocks defined by Hoechst and N-banding techniques. The results of this analysis
indicate a good correlation between the N-banded regions and 1.705 repeats and between the
Hoechst-bright regions and the 1.672 repeats. However, the molecular basis for banding does not
appear to depend
exclusivelyon DNA content, since heterochromatic blocks showingidentical banding
patterns often contain different combinations of satellite repeats. The distribution of satellite repeats
has also been analyzed with respect to the male fertility factors of the Y chromosome. Both loopforming (kl-5, kl-3 and ks-I) and non-loopforming (kl-2 and b - 2 ) fertility genes contain substantial
amounts of satellite DNAs. Moreover, each fertility region is characterized by a specific combination
of satellite sequences rather than by an homogeneous array of a single type of repeat. We discuss the
possible functional role of these satellite sequences in the light of the available information on the Y
chromosome fertility factors of D.melanogaster.
T
HE Y chromosome of Drosophila melanogaster is
an entirely heterochromatic element which carries a limited but well defined set of genetic functions:
the fertility factors, a series of loci whichare essential
for male fertility (BRIDGES1916; STERN1929; BROSSEAU 1960); the bobbed locus (bb), which is allelic to
the bobbed locus located inthe X heterochromatin and
corresponds to the ribosomal cistrons (for review see
RITOSSA1976); a pairing site(collochore) required
during male meiosis for regular pairing and segregation of the sex chromosomes (COOPER1964); a locus
necessary for normal male meiosis, whose deficiency
causes the appearance of needle shaped crystals inthe
primary spermatocytesand anabnormal chromosome
distribution at meiosis (HARDYet al. 1984; LIVAK
1984, 1990).
Classical genetic and cytological analyses of these
functions have beenfor a long time precluded because
the Y chromosome does not undergo meiotic recombination and is included in the chromocenter of polytene chromosomes.More recently, by applying a
series of banding techniques to neuroblast prometaphase chromosomes, it was possible to differentiate
Genetics 1 2 9 177-189 (September, 1991)
the mitotic Y chromosome into 25 regions (GATTIand
PIMPINELLI1983). Thusan extensivecytogenetic
analysis could be undertaken which permitted the
mapping of the functions carried by the Y chromosome and provided some insights into their genetic
organization (GATTI and PIMPINELLI
1983; BONACCORSI et al. 1988).
The most extensively studied Y chromosome functions are the fertility factors. Four fertility factors
have been cytologically localized on the long arm of
the Y chromosome and two on the short arm(KENNISON 1981; HAZELRIGG,
FORNILIand KAUFMAN 1982;
GATTIand PIMPINELLI
1983). Starting from the tip of
the long arm they are designated as k1-5, kl-3, kl-2, kl1, ks-1 and ks-2; the existence of kl-4, postulated by
BROSSEAU
(1960), has not been confirmed (KENNISON
198 1; HAZELRICG,FORNILIand KAUFMAN 1982;
GATTIand PIMPINELLI
1983). Moreover, it has been
shown that each of k1-5, kl-3 and KS-1 fertility factors
possesses extremely large physicaldimensions;they
are defined by a series of noncomplementing sterile
breakpoints scattered over chromosome regions containing up to 4000 kb of DNA. The cytogenetic data
178
S. Bonaccorsi and A. Lohe
on kl-2, kl-1 and ks-2 enabled a ratherprecise localization of these loci along the Y but not an estimation of
their physicalsize (GATTI and PIMPINELLI1983; C .
PISANO,S. BONACCORSI
and M. GATTI,manuscript in
preparation). More recently, it has been shown that
the kl-5, kl-3 and ks-I fertility factors are responsible
for the development of three giant lampbrush-like
loops in primaryspermatocyte nuclei. These loops are
analogous to those described in Drosophila hydei and
are thought to represent the cytological manifestation
of fertility factor activity during this stage of spermaet al. 1988).
togenesis (BONACCORSI
A number of studies performed during the past
yearshaveshown that a seriesofsimplesequence
satellite DNAs are located on the Y which together
et al.
account for about 80% of its length (PEACOCK
1977; APPELSand PEACOCK1978; STEFFENSEN,
APPELS and PEACOCK1981). D. melanoguster contains
four well defined satellite DNAs which amount to
about 20% of the genome and can be divided into
two groups based on sequencecomplexity (GALL,
et al. 1973). One
COHENand POLAN197 1; PEACOCK
group contains tandem repeats of a simple sequence,
only 5, 7 or 10 bp in length; these satellites band at
1.672, 1.686 and 1.705 in CsCI. The other group is
represented by the 1.688 satellitewhichconsistsof
tandem repeats of a longer sequence, 359 bp in length.
Molecular cloning of D. melanogaster satellite DNA
provided a more refined view of this genetic material,
by showing that more than one sequence is present in
each satellite (LOHEand BRUTLAC1986). Together,
11 related sequences were identified in thethree
simple satelliteDNAs, all mapping to the heterochromatic portions of the genome, and 9 of them were
shown to be present on the Y chromosome (LOHEand
and ROBERTS,unpubRoberts 1988; LOHE,HILLIKER
lished). However, the in situ hybridization experiments of LOHEand ROBERTS(1988) were performed
on wild-type mitotic chromosomesand did not permit
a precise localization of the satellite sequences with
respect to the cytological and genetic entities previ1983;
ously identified on theY (GATTIand PIMPINELLI
BONACCORSI
et al. 1988).
Here we describe a series of in situ hybridization
experiments performed on a number of cytologically
determined rearrangements involving the Y chromosome. The heterochromatic breakpoints of these rearrangements provided an array of unambiguous
landmarks along the Y which have been used to localizeeachsequence
onthe various heterochromatic
blocks defined by the banding techniques. This allowed us to relate the molecular composition of specific Y chromosome blocks to their cytochemical and
functional properties.
MATERIALS AND METHODS
Drosophila stocks All the Y autosome translocations used
here were generated by LINDSLEY
et al. (1972) using a B"Yy+.
The genetics and cytology ofthese translocations have been
described by LINDSLEY
et al. (1 972), GATTIand PIMPINELLI
(1 983) and by BONACCORSI
et al. (1 988).
All the T(X;Y)s used here, except T(X;Y)Tl3, are fertile
reciprocal translocations having the X breakpoint in the
proximal heterochromatin; they involve an X chromosome
marked withy wfand aE'Yy' chromosome (KENNISON198 1;
HARDY
et al. 1984). T(X;Y)Tl3 is a sterile reciprocal translocation (KENNISON198 1) having the Y breakpoint in region
h5 and carrying an additional deficiency that encompasses
regions h5-h6. The X and Y chromosome breakpoints of
T(X;Y)FIZ, T(X;Y)F15 and T(X;Y)T13 were determined in the
present study and are reported in Table 1 , which summarizes the cytological features of all the rearrangements used
here.
Before being used for in situ hybridization experiments,
all the Y chromosome rearrangements were reexamined
cytologically by Hoechst and N-banding according to GATTI
and PIMPINELLI
(1 983). This analysis revealed that 17 of the
18 rearrangements previously described (GATTIand PIMPINELLI 1983; HARDYet d . 1984; BONACCORSI
et d . 1988)
maintained their original constitution. T(Y;Z)A77 picked up
an additional deficiency encompassingregions h 1-h 16.
All the othermutations, special chromosomes and genetic
markers used in this work are described by LINDSLEY
and
GRELL(1968). All stocks and crosses were maintained at
25" f 1 " .
Recombinant plasmids:Isolation and cloning of D. melanogaster satellite DNA sequences are described in detail
elsewhere (LOHEand BRUTLAC1986). The eight satellite
DNA clones used inthe present work are reported in Table
2. The 1.672-1 plasmid clone contains a tandem array of
AATAC satellite repeats adjacent to part of a mobile element DNA (LOHEand BRUTLAC1987b). AnEcoRI fragment of this plasmid contains the complete satellite array of
181 bp flanked by 7 1 bp of mobile element DNA. This
fragment was subcloned intothe pSP65 vector and the
plasmid was named 1.672-18 1 .
['HIRNA probes of each sequence were synthetized from
pSP64/65 DNA templates. Nick-translated ['HIDNA
probes were also synthesizedand used for asubset of in situ
experiments with 1.672-1 8 1 , 1.705-42 and 1.705--34.
Stringency of hybridization: The mean melting temperature (Tm) of the simple satellite DNAs is dependent on
nucleotide composition and sequence, and varies widely
among these closely related sequences (LOHEand BRUTLAG
1987a). Since different satellite repeats can show up to 80%
sequence homology, stringent hybridization criteria were
established to avoid cross-hybridizationamong the classes of
satellite sequences. The Tm values for RNA-DNA hybrids
were determined for each of the eight satellite probes and
a hybridization temperature of 10-1 3" below the Tm value
was chosen for the in situ hybridizations (Table 2).
I n situ hybridization: Mitotic chromosome preparations
were made following the proceduredescribed by GATTIand
PIMPINELLI
(1 983). Minor modifications to their procedure
were that slides have been frozen in liquid nitrogen and,
after removing the coverslip, immediately placed in 95%
ethanol and air dried.
The in situ hybridization procedure is similar to that used
et al. (1 977). Freshly made, air driedslides were
by PEACOCK
placed in 0.2 M HC1 (37") for 30 min, rinsed briefly in
distilled water, dehydrated in 70% ethanol (2 times), 95%
ethanol (2 times) and air dried. 3H-labeled probe (1 X lo5
cpm) in 3 X SSC, 50% formamide (vol/vol) was applied to
the slide, an 18 X 18 mm coverslip placedover the solution,
and the coverslip was sealed with rubber cement. Prior to
incubation slides were placed at 65" in an air incubator for
15 min and slides were then placed at the hybridization
Y Chromosome SatelliteDNAs
179
TABLE 1
Cytological features of rearrangements
Y breakpoint"
Translocation
Other breakpoint Reference'
28B
85E
48E
h30
h26
34B
38B
h26
33B
72AB
36C
h29
76D
h26
26B
25A
24D
31EF
35CD
66A
57F
Fertilitf
1,
hl/h2
TCy;2)R50
1 , 3,
h2/h3
T(Y;3)G42
1, 3, 5
h3
TCy;2)D 19
2, 3,
h4
T(X;y)V24
2
h5 Df(h5-h6)
T(X;Y)T13
1, 3, 5
h7 ln(h1-h3)
TCy; 2)G
74
1, 3, 5
h9/h 10
TCy;2)P5 7
F
2, 3, 4
hl1
T(X;Y)E 1
F
1, 3
hll
TCy;2)E92
F3
1,
h13
TCy;3)B223
S(R1-1)
1, 3
h14
T(Y;2)B242
2
F
h15
T(X;Y)F1 2
F
1, 3
h15
TCy;3)B115
2
F
h15/h16
T(X;y)Fl5
S(kZ-2,3,5)
3
1,
h16 Df(h3-h13)
T(Y;2)H121
h19 Df(h1-hl6)
S(KL)
3
1,
TCy; 2)A
77
h2 1
5
S ( h - 1 ) 3,
1,
T(Y;2)PS
S(ks-1)
1, 3, 5
h2 1
TCy; 2)A
162
5 S(h-1)
3,
1,
h2 1
T(Y;2)J165
S(kS-1)
1 , 3, 5
h22/h23
T(Y;3)R119
5
S ( h - 1 ) 3,
1,
h23/h24
T(y;2)J163
S. BONACCORSI,DIMITRI
P.
and M.GATTI in
The nomenclature of the X chromosome breakpoints is described in detail in S. PIMPINELLI,
preparation. The X heterochromatin has been subdivided into 9 regions; region h29 corresponds to the nucleolus organizer and region h33
to thecentromeric area.
A slash between two Y regions means that the rearrangement is broken at thejunction of these two regions.
* The fertility factors which are disrupted by the rearrangement are indicated between the brackets.
1 , LINDSLEY,
et al. (1972); 2, KENNISON(1981); 3, GATTIand PIMPINELLI
(1983); 4, HARDY
et al. (1984); 5, BONACCORSI
et al. (1988).
+
+
+
+
Mean melting temperatures of satellite RNA-DNA hybrids and
hybridization temperatures
1.705-42
1.705-34
1.686-198
1.672-38
1.672-349
1.672-181
1.672-453
1.672-563
Repeating
sequence
AAGAG
AAGAGAG
AAGAC
AATAT
AATAG
AATAC
AATAAAC
AATAGAC
Tm value of
RNA-DNA ("C)
Hybridization
temperature ("C)
59
48
50
40
16
23
23
25
35
61
53
26
35
36
37
48
S(k1-5) 3,
S(R1-5)
5
S(kZ-5)
F
5
S(k1-3)
S(kl-3,5)
S(R1-3)
RESULTS
TABLE 2
Satellite
clone
5
The melting temperatures were determined by P. A. ROBERTS
and A. R. LOHE(unpublished results).
temperature (see above) for 16 hr. Following hybridization,
coverslips were removed and residual probe washed away
by incubation in the hybridization solution at the hybridization temperature for15 min (2 times). Slides were washed
in 2 X SSC for 15 min, treated with RNAse A (2 rg/ml in
2 X SSC, 30 min at room temperature), washed again in 2
X SSC (4 times, 15 min each), placed in 70% ethanol, 95%
ethanol (2 times), airdried, dipped in Ilford K2 emulsion
(diluted 1:1 with water) and exposed at 4 " in a light-tight
box. Exposure times were determined empirically andvaried for different probes from
3 days to several weeks. Slides
were developed in Kodak Dektol D l 9 (2 min.), washed in
water and fixed in Kodak Fixer for4 min. They werestained
for 20 min. with a 10%solution of freshly preparedGiemsa
(BDH, R66) in 8 mM KHzP04, 6 mM Na2HP04,pH 6.8,
rinsed in tap water,air dried and mountedin immersion oil
(Zeiss). Slides were stored indefinitely in this way.
I n situ hybridization experiments were performed
using eight satellite DNA clones recently isolated by
LOHEand BRUTLAG
(1986). The repeating sequence
contained in each clone, together with the hybridization temperatures used for thein situ experiments are
reported in Table 2. Four sequences (AAGAG, AAGAGAG, AAGAC, AATAT) map to multiple sites
along the Y and to other heterochromaticportions of
the Drosophila genome; three sequences (AATAC,
AATAAAC, AATAGAC) map only to unique sites
on the Y, and one (AATAG) toa single site on the Y
and to the 2nd chromosome heterochromatin (LOHE
and ROBERTS1988).
To finely map the 8 satellite sequences associated
with the Y chromosome we used 16 Y autosome and 5
X-Y cytologically determined translocations (GATTI
and PIMPINELLI
1983; BONACCORSI
et al. 1988) whose
breakpoints span the length of the Y chromosome.
The breakpoints of these translocations are diagramatically reported in Figure 1. All these rearrangements were induced on a FYy+ which carries
two X heterochromatin blocks appended at the distal
ends of YL and P,respectively (GATTIand PIMPINELLI
1983). The presence of these blocks of X heterochromatin does not interfere with the satellite mapping
along the Y chromosome because their principal component is the 359-bp repeats of the 1.688 satellite
DNA, which are absent from the wild type Y (HILLI-
S. Bonaccorsi and A. Lohe
180
G74
mo
-
(AAGAG)
Lu
u
w
(AAGAGAG),
(AAGAC),
(AATAT),
PA
P7AO
u
U
u
U
(AATAG),
u
U
U
U
(AATAC),
U
(AATAAAC),
U
(AATAGAC),
U
1
2
35 4
A
I
6 7 6
9 11
10
13
12
E
14 16
15
17
16 19
22
21
23 24 25
C
FIGURE1.-Localization of 8 satellite DNA sequences along the Y chromosome of Drosophila melanoguster. The diagram is a schematic
representation of the Y chromosome stained with Hoechst 33258 (GATTI and PIMPINELLI
1983). The chromosome is subdivided into 25
regions defined by the degree of fluorescence and the presence of constrictions. Filled segments indicate bright fluorescence, hatched
segments indicate dull fluorescence and open segments indicate no fluorescence. C: centromere; N: N-banded regions. The vertical lines
above the diagram indicate the Y chromosome breakpoints of the 21 translocations used for the in situ hybridization experiments. G 7 4 , 8 2 4 2
and P8 were used for the gross localization of each satellite sequence, the other translocations were employed for fine mapping (see text and
Table 1 for further explanation). The horizontal lines below the upper diagram indicate the localization of each sequence. The breakpoints
that allowed each localization are reported as short vertical lines distributed along each horizontal line. Each sequence has been localized in
chromosome regions defined by at least two breakpoints; the only exception is represented by the localization of the AAGAG repeats within
region h24-h25 (see text). The thin horizontal lines below the lower diagram indicate the maximum physical size of the fertility factors and
1983; BONACCORSI
et al. 1988). In the lower row, thick
the thicker lines below them their minimum physical size (GATTIand PIMPINELLI
lines indicate the loop forming regions (BONACCORSI
et al. 1988).
KER and APPELS 1982). T h e X heterochromatin also
contains low amounts of AACAG and AATAT repeats but these sequences are restricted to the periand
centromeric areaof the X chromosome (HILLIKER
APPELS1982; LOHEand ROBERTS1988). None of the
X;Y translocations used in the present work has an X
breakpoint falling near the centromere(see Table 1);
thus, X-Y translocations could be used to map these
repeats along the Y chromosome without significant
interference from the centric materialof the X .
For each sequence, agross localization was obtained
by using three Y autosome translocations [T(Y;2)G74,
T(Y;2)B242 and T(Y;2)P8] whose breakpoints approximately divide the Y chromosome intofour equal
sections (see Figure 1). Once its overall distribution
was determined, each sequence was finely mapped by
using a series of breakpoints that further subdivide
the Y. In general, each sequence was mapped to chromosome regions defined by two breakpoints. In some
cases, the examination of particularly elongated prometaphasechromosomespermittedamore
precise
localization of a sequence within these regions. Examples of hybridization on complete metaphases carrying Y autosome translocations are shown in Figure
2. Figure 3 shows the cytological definition of the
breakpoints of three Y chromosome rearrangements
by sequential staining with Hoechst 33258 (Figure 3,
a-c) and N-banding (Figure 3, d-f) and illustrates the
method used for the localization of a sequence along
a Y region definedby three translocation breakpoints.
Localization of AAGAG repeats: T h e AAGAG
sequence is the main component of the 1.705 satellite
DNA; it is very abundant, accounting for about 5.6%
of the haploid genome of D.melunoguster (LOHEand
1986). This sequence maps to multiple sites
BRUTLAG
along the Y and, in different amounts, to the heterochromatin of all the chromosomes of the complement
1981; LOHEand
(STEFFENSEN,
APPELSand PEACOCK
ROBERTS1988; see also Figure 2).
In situ hybridization on T(Y;2)G74 shows the presence of labeling on both the Y-distal and Y-proximal
elements of this translocation. The Y-distal element of
G74 exhibits a single cluster of silver grains near the
translocation breakpoint while its Y-proximal element
exhibits grainson two sites, one close but notadjacent
to the breakpoint, and the other on the short arm.
Silver grains are also present on both sides of the
breakpoint of T(Y;2)B242 (Figure 4c). However, in
this case it is the Y-distal element which exhibits two
clusters of grains, one located near the breakpoint
and the other in the distal part of the long arm. The
Y-proximal element of B242 appears to be heavily
Y Chromosome Satellite DNAs
labeled only on the short arm, showing the chromosome segment h15-h20, comprised between the
breakpoint and the distal end of the nucleolar constriction, completely devoid of grains. Accordingly,
the Y-proximal element of TCy;2)P8 is labeled onlyon
the long arm, while its Y-distalelement shows a heavy
labeling throughout its length. Together these data
indicate that the labeling is present only on three of
the segments defined by G74, B242and P8;the B242P8 segment, which comprises regions h15-h20, does
not appear to contain AAGAG sequences.
The remaining three segments of the Y, each showing at least one main site of hybridization with this
sequence, have been further dissected using a series
of suitable breakpoints. The Y breakpoints used and
the results of this analysisare reported below (see also
Figure 1).
No AAGAG repeats are present in regions hl-h2
as shown from the lack oflabeling on the Y-distal
elements of both TCy;2)R50 , and T(Y;3)G42 (Figure
3g). A heavy labeling is present on the Y-distal elements of both TCy;2)D19 (Figure 3h) and T(X;y)V24
(Figure 3i), indicating that the AAGAG repeats are
located in region h3.
The Y-proximal element of V24 exhibits a heavy
labeling site quite close to its breakpoint (Figure 3i);
the Y-distal element of G74 also showsa heavy labeling
site close to its breakpoint. Together these observations demonstrate the presence of the AAGAG sequences somewhere within regions h4-h6.
Neither the Y-proximal element of G74 nor the Ydistal element of TCy;2)P57 exhibit labeling close to
the breakpoint; this provides evidence that no AAGAG sequences are located in regions h7-h9. The Yproximal element of P57, on the other hand,appears
to be heavily labeled just at its breakpoint. Labeling
is alsoobservedclose to and on both sidesof the
breakpoints of T(X;y)El and T(Y;2)B92 (Figure 4a).
The Y-distal element of Tfy;3)B223 is labeled close to
the breakpoint, but no consistent labeling is apparent
near thebreakpoint of the Y-proximal element, which
carries regions h14-h25. Thus, AAGAG repeats appear to be located in region h l O-h13 but no clear
evidence of their presence in region h14 has been
obtained. These results also indicate that theAAGAG
sequence is located in region h 10,but do not permit
a fine mapping of this sequence within region h l 1h13. The data are consistent with a localization ofthe
AAGAG sequence in region h l 1 or h l 2 or both.
As mentioned above, no AAGAG repeats are present in the Y chromosome segment comprised between
the breakpoints of B242 and P8,while multiple sites
of labeling are observed along the remaining portion
of the shortarm (regions h21-h25). The breakpoints
of T(Y;2)P8, T(Y;2)A162(Figure 2a) and T(Y;2)J165 all
fall in region h21, in the proximal, middle and distal
third, respectively. The Y-distal elements ofthese
181
translocations carry regions h2 1-h25 and are heavily
labeled in each case. The Y-proximal element of P8 is
not labeled whilethe proximal elements of both A1 62
and J165 showheavylabeling,
indicating that the
AAGAG sequence is localized in region h2 1.
The Y-distal element of J165 shows a heavy labeling
sitewhich,in
particularly elongated chromosomes,
appears to be separated from its breakpoint and from
an additional site of labeling located more distally on
the short arm. Twosites of labeling are still apparent
on the Y-distal element of T(Y;3)R119 (Figure 4e),
whereas onlyone site of labeling is left on the Y-distal
element of TCy;2)J163 (Figure 4g). Thus, tandem repeats of the AAGAG sequence are present in region
h23 and in an additional site comprised between the
middle of region h24 and the end of the short arm.
Since no breakpoints are available along region h24h25 a more refined localization of the AAGAG sequence within this interval was precluded. In conclusion, AAGAG repeats were mapped to five separate
sites along the Y, corresponding to regions h3-h6 and
h O-h
1 1 3
of the long arm and to
regions h2 1, h23 and
h24-h25 of the short arm.
Localization of AAGAGAG repeats: The 7-bp sequence AAGAGAG is a minor component of the
1.705 satellite DNA. Repeats of this sequence, which
are located on the Y chromosome and on the heterochromatin of the second andthird chromosomes,
account for about 1.5% of the haploid genome of D.
melunoguster (LOHE and BRUTLAC
1986; LOHE and
ROBERTS1988). I n situ hybridizationanalysisusing
G74, B242 (Figure 4d) and P8 showed the presence
of two main sitesof hybridization of the AAGAGAG
sequence along the Y chromosome. One site is located
on the long arm, distal to the breakpoint of G74, and
the other on the short arm, distal to the breakpoint
of P8.
The use of four additional breakpoints on the long
arm and four on the short demonstrated
arm
a partial
coincidence betweenthe location of this sequenceand
that of the AAGAG repeats. The labeling pattern of
R50, G42, Dl9 and V24 demonstrated that AAGAGAG repeats are restricted to region h3 of the long
arm and do not extend into regions hl-h2 or h4-h6.
In addition, none of the Y-proximal elements of P8,
A162, J165, R119 (Figure 4f) and J163 showed a
consistent labeling at the breakpoints, whereas all the
Y-distal elements ofthese translocations are clearly
labeled. The AAGAGAG cluster of the short arm
appears therefore to map between the breakpoint of
J16.3, at the junction between region h23 and h24,
and the distal end of the short arm.
Localization of AAGAC repeats: The AAGAC
sequence is a component of the 1.686 satellite and
comprises about 2.4% of the haploid genome. It maps
to multiple sites along the Y and to the heterochromatin of the second chromosome (LOHEand BRUTLAG
182
S. Bonaccorsi and A. Lohe
-
FIGURE2.-Examples of in situ hybridization on metaphase plates carrying different Y autosome translocations. The numbers identify
each chromosome of the complement and the arrows point to the translocation breakpoints. YP and YD indicate the Y-proximal and Y-distal
elements of each translocation. a: T(Y;2)A162, translocation breakpoints in region h21 of the Y chromosome and in 2L (31EF), hybridized
with the AAGAG probe. The heterochromatin of all the chromosomes appears heavily labeled. In particular, the Y-proximal element shows
a heavy signal at the translocation breakpoint and multiple sites of labeling on the long arm; the Y-distal element shows a heavy labeling on
Y Chromosome Satellite DNAs
1986; LOHEand ROBERTS1988). Repeats of this sequence are present in the regions distal to the breakpoints of G74 (hl-h7) and P8 (h21-h25) and in the
interval comprised between the breakpoints ofG74
and B242, (regions h7-hl5) but not in the segment
defined by the breakpoints of B242 and P8, corresponding to regions h 15-h2 1.
For the fine mapping of the AAGAC sequence
within these regions 11 additional breakpoints were
used. The hybridization patterns on R50, G42, Dl9
and V24 demonstrate that the AAGAC repeats are
absent from regions hl-h2, abundantly present
throughout region h3 and scarcely present within
regions h4-h5. The results obtained using P57, B92
(Figure 4b) and E l clearly indicate that the AAGAC
repeats present in the middleof the long arm are
restricted to region hl0. Finally, the labeling patterns
on P8, A162 and J165 and J163 indicate the presence
of two hybridization sites on the short arm: one in
region h21 and the other between the breakpoint of
J163 (Figure 4h) and the terminus of the short arm,
somewhere within region h24-h25.
Localization of AATAT repeats: The AATAT
repeats represent the major component of the 1.672
satellite DNA. They account for 3% of the haploid
genome and map in the heterochromatic portions of
all the chromosomes of the complement with a principal localization in the 4 and the Y (LOHEand BRUTLAG 1986; LOHE and ROBERTS1988). AATAT repeats are contained in all the four segments defined
by the breakpoints ofG74,B242 and P8. Further
analysis of the location of the A A T A T repeats was
carried out using 10 additional breakpoints on the
long arm and two on the short arm.The
hybridization
patterns observed on R50 (Figure 5a) and G42
strongly suggest that region hl-h2 contains the AAT A T repeats, while region h-3 is devoid of these
sequences. The whole region comprised between the
breakpoints of V24 and P57 appears heavily labeled
with the AATATsequence. Moreover, T(X;Y)T13 and
G74 (Figure 5b) both exhibit a quite strong signal on
each sideof their breakpoints, indicating thatthe
AATAT sequences are located in region h4 and in
regions h7-h9.However,they
do not prove their
localization in region h5-h6, since TI3 carries a deletion of this region, and do not permit a decision on
whether they map throughout the entire h7-h9 region or only to aportion of it.
183
Neither the Y-proximal element of P57 nor the Ydistal element of B242 show any labeling located close
to their breakpoints (Figure 5c). Thus, region h10h14 does not appear to contain A A T A T sequences.
The Y-proximal elements ofB242 (Figure 5c) and
T(X;Y)F12 show a heavy labeling close to their breakpoints; the Y-proximal elements of both T(Y;3)B115
and T(X;Y)Fl5 are slightly labeledat their breakpoints,
while no grains are observed near the breakpoint of
the Y-proximal element ofT(Y;2)H121. Conversely,
the Y-distal elements of these translocations consistently show silvergrains at their breakpoints. Together
these observations strongly suggest that AATAT sequences are mainly clustered within region h15, still
present in region h16 and absent from regions h 17
and h18.
The Y-distal element of T(Y;2)A77 and the Y-proximal elements of P8, J165 and J163 (Figure 5d) donot
exhibit any labelingnear their breakpoints, indicating
that noAATAT sequences are located in regions
h19-h23. On the otherhand, the Y-distal elements of
P8, J165 and J163 (Figure 5d) show a rather heavy
labeling suggesting that the localization of A A T A T
repeats on the short armis distal to the breakpoint of
J163, somewhere within region h24-h25.
Localization of AATAG,AATAC,AATAAAC
and AATAGAC repeats: All these sequences were
clonedas minor components of the 1.672 satellite
DNA (LOHEand BRUTLAG1986), each amounts to
only 0.1-0.5% of the D. melunoguster genome (LOHE
and BRUTLAC1986) and maps to unique sites along
the Y chromosome (A. R . LOHE, A. J. HILLIKER
and
P. A. ROBERTS unpublished). Tandem arrays of the
AATAG repeats are mainly located at a single site of
the Y chromosome and on the heterochromatin of
chromosome 2 (LOHE and ROBERTS 1988). The
AATAG sequence hybridizes only with
the Y-distal
element of G74 and with the Y-proximal element of
V24 and is thus restricted to region h4-h7. A more
precise localization of the AATAG sequence within
this region was obtained using T13. Only the Y-distal
element of thistranslocation, carrying regions hl-h4,
is labeledwith the AATAG probe, mappingthese
repeats to region h4. However, since the Y-proximal
element of T13 carries a deficiency that encompasses
regions h5-h6, we cannot exclude that some AATAG
sequences are also present in the deleted region.
The AATAC sequence is exclusively located onthe
both the second chromosome heterochromatin and the Y short arm. b and c: T(Y;2)G74, breakpoints in region h7 of the Y chromosome and
in 2L (34B), hybridized with the A A T A T (b) and the AATAC sequences (c). In b, the Y-proximal element of this translocation appears
heavily labeled at the translocation breakpoint and atadditional sites on both the long and the short arm; the Y-distal element shows a strong
signal extending from the translocation breakpoint to most of the long arm. Note the heavy labeling of the fourth chromosomes and of the
tip of the attached X s . The pericentromeric area of the third chromosome is also labeled while the second chromosome heterochromatin
appears completely devoid of grains. In c, after hybridization with the AATAC probe only the attached-XY and the Y-distal element of the
translocation are labeled at a site located on the long arm, distal to the breakpoint of G74. Thus thehybridization is restricted to a single site
of Y'..
184
S. Bonaccorsi and A. Lohe
FIGURE3.-Mapping the AAGAG sequence to region h3. The six upper panels (a-f) show the reciprocal elements of three translocations
sequentially stained with Hoechst 33258 (a-c) and N-banding (d-f). The three lower panels show the same translocations after in situ
hybridbation with the AAGAG sequence (g-i). T h e arrows point to thetranslocation breakpoints and the numbers alongthe Y chromosome
1983). a, d, g: T(Y;3)C42, breakpoint at the distal end of region h3
indicate cytological landmarks (cf. Figure 1 and GATTIand PIMPINELLI
(a, d). The Ydistal element shows hybridization only on the third chromosome heterochromatin (g, left), while a heavy signal is observed at
the translocation breakpoint and atmultiple sites of the Y-proximal element (g, right). b, e, h: T(Y;2)D19, breakpoint in the middle of region
h3 @, e). The Ydistal element shows a site of hybridbation coincident with the translocation breakpoint (arrow) and an area of heavy labeling
on thesecond chromosome heterochromatin (h, left). T h e Y-proximal element is labeled at thetranslocation breakpoint, in the middle of the
long arm and on the short arm (h, right). c, f, i: T(X;y)V24, breakpoint between regions h3 and h4 (c, f). The Ydistal element is labeled at
the translocation breakpoint (i. left). A weaker signal is observed near the translocation breakpoint of the Y-proximal element which also
shows two additional sites of hybridi~ationin the middle of the long arm andon the shortarm (i, right).
Y chromosome (LOHEand ROBERTS1988).It maps to
a single site located between the breakpoints of V24
and G74 (Figure 2c). None of the reciprocal elements
of T13 is labeled with this sequence. Thus, the AATAC repeats map to regions h5-h6 which, as mentioned above, are deleted in T13. The AATAAAC
repeats map to region h22, in the interval comprised
between the breakpoints of J165 and RI 19, while the
AATAGAC sequence is localized in region h10, between the breakpoints of E l and P57.
DISCUSSION
In the present study we have finely localized eight
satellite DNAsequences along the Y chromosome.
Several improvements in the satellite mapping strategy compared with previousstudies were essential for
Satellite
Y Chromosome
185
DNAs
a
b
d
VL
w
f
VL
4
FIGURE4.-h situ hybridbation of the AAGAG (a, c, e, g), AAGAC (b, h) and AAGAGAG (d, f) sequences on different Y autosome
translocations. The arrows point to the translocation breakpoints. a, b: T(Y;2)R92,breakpoint in region h 1 1 (see Fig. I for the localization of
this and the following breakpoints), hybridized with the AAGAG (a) and AAGAC (b) sequences. The Y-distal element shows the same pattern
of labeling after hybridization with either of these sequences (a and b, left). The Y-proximal element of this translocation is labeled at its
translocation breakpoint after hybridi~ationwith the AAGAG sequence (a, right) but not after hybridization with the AAGAC sequence (b,
right). c, d: T(Y;2)B242, breakpoint in region h14, hybridized with the AAGAG (c) and AAGAGAG (d) sequences. In (c, left) the Y-distal
element of this translocation shows a site of heavy labeling at the translocation breakpoint and a second site of hvbridization located more
distally. In (d. left) only a distal site of hybridi~ationis observed, while no grains are present at the translocation breakpoint. Note that the Yproximal element of this translocation shows a heavier labeling of the short arm with the AAGAG (c, right) than with the AAGACAG (d,
right) sequence. e, f T(Y;3)R119, breakpoint at the proximal end of region t123, after hvbridi7~tionwith the AAGAG (e) and AAGAGAG
(f) sequences. In(e) the Y-proximal element (left) shows a clear site of labeling at the translocation breakpoint, distal to thenucleolus organizer
constriction (NO), and multiple sites of labeling on the long arm, while the Y-distal element (right) shows two close but distinct sites of
labeling. In (f) only a distal region of the Y-proximal element (left) is labeled, and a single site of hybridization is present on the Y-distal
element (right). g, h: T(Y;2)J163,breakpoint at the distal end of h23, afterhybridbation with the AAGAG (g) and AAGAC (h) sequences. In
both cases the Y-proximal element shows a rather heavy labeling close to the translocation breakpoint and multiple sites of hybridization on
the long arm (g and h, left), while the Y-distal element appears labeled at a stngle site close to the translocation breakpoint (g and h. right).
Note the heavier labeling of the second chromosome heterochromatin observed after hybridization with the AAGAG sequence (g. right).
186
S. Bonaccorsi and A. Lohe
4
.
0
a
b
FIGURE5.-In sifuhybridization of the AATATsequence on different Y-2 translocations. The arrows point to thetranslocation breakpoints.
a: T(Y;2)R5O, breakpoint in region h2. The Ydistal element (left) shows a site of labeling close to the translocation breakpoint while the Yproximal element (right) shows multiple sites of hybridization both on the long and on the shortarm. b T(Y;2)G74, breakpoint in region h7.
T h e Y-distal element (left) is labeled at its translocation breakpoint and at an additional site located more distally. The Y-proximal element
(right) is labeled at the translocation breakpoint, near the centromereregion and on the short arm. c: T(Y;2)B242, breakpoint in region h14.
T h e Ydistal element (left) shows a heavy labeling only on its distal portion, while the Y-proximal element shows an area of heavy labeling
comprised between the translocation breakpoint and thenucleolar constriction (NO) and an additional site of hybridbation on the short arm.
d: T(Y;2)J163, breakpoint at the distal end of region h23. T h e Y-proximal element (left) does not show any labeling along the entire region
comprised between the translocation breakpoint and the nucleolar constriction (NO), while two main sites of hybridization are evident near
the centromere and in a rather distal portion of the long arm. The Ydistal element (right) is clearly labeled right at the translocation
breakpoint.
the generation of a high resolution mapof the Ylocatedsatellites. First, w e usedcloned probes that
contain a single sequence repeated in tandem for the
entire length (LOHEand BRUTLAC1986) ratherthan
probes from gradient-purified, bulk satellites.Second,
different satellites wereaccurately mapped using stringent hybridization conditions that are specific for each
of the satellite repeats, to avoid cross hybridization
betweentheseclosely related sequences (LOHEand
BRUTLAC1987a). Finally, we mapped the satellite
sequences with respect to translocation breakpoints.
Using 16 Y autosome and 5 X-Y cytologically determinedtranslocations(GATTI and PIMPINELLI
1983;
BONACCORSI
et al. 1988)a ratherfine mapping of each
sequence was possibleevenin
the absence of extremely elongated chromosomes, since for every
translocation analyzed witha given probe we only had
to determine on which side of the breakpoint the
hybridization occurred. In this way most of the satellitesequencescouldbeunambiguouslyassigned
to
one or more heterochromatic blocks. This permits us
to relate the molecular composition of the Y regions
to their banding patterns and their functional properties.
Relationship between satellite sequence distribu-
tion and chromosome banding: Three types of regions can be basically distinguished along the Y after
staining with quinacrine, Hoechst and N-banding (6
with the diagram in Figure 1): (1) Hoechst and quinacrine brightly fluorescent and N-banding negative
regions (hl, h2, h4, h6, h8, h9, h15, h17, h22 and
h24); (2) Hoechst and quinacrine dully fluorescent
and N-banding negative regions (h7, h l 1, h13, h19
and h20); (3) Hoechst and quinacrine nonfluorescent
and N-banding positive regions (h3, h5, h10, h12,
h14, h21, h23 and h25). It has been suggested that
this cytochemical heterogeneity reflects the different
base composition of the highly repeated DNAs contained in the three classes of regions. In particular,
fluorochrome-bright blocks have been associated with
the 1.672 AT-rich satellite DNA while N-banded regions have been suggested to co-map with the relatively GC-rich 1.705 satelliteDNA (GATTI,PIMPINELLI and SANTINI1978; PIMPINELLI,
SANTINIand
GATTI1978; APPELSand PEACOCK
1978; STEFFENSEN,
APPELS and PEACOCK
1981; GATTIand PIMPINELLI
1983).
The high resolution mapof the Y chromosome
satellite DNA sequences reported here shows that in
somecases a given sequence could be mapped to
Y Chromosome Satellite DNAs
homogeneously stained blocks defined by 2 or 3
breakpoints, while inothers, due to the
lack ofsuitable
breakpoints, it was mapped to asegment composed of
two differently stained blocks(see Figure 1). The
results obtained on the homogeneously stained regions clearly show that the AT-rich sequences from
the 1.672 satellite DNA are almost exclusively localizedinHoechst bright regions while the N-banded
regions always contain one or more sequences from
the relatively GC-rich1.705 and 1.686 satellite DNAs.
The only exception is represented by region h16, that
is Hoechstnegative and appears to accommodate
some A A T A T repeats. In this respectit must benoted
that theresolution of our cytological method does not
permit us to exclude that the breakpoint of F15 falls
in the very proximal part of region h15; the Y-proximal element of this translocation could carry a very
small proportion of the Hoechst bright region h15,
not detectable by banding methods, which could be
responsible for the labeling observed after hybridization with the AATAT sequence.
Based on these results we can make someinferences
about the localizationof satellite DNAsin regions
which contain differently stained heterochromatic
blocks. For example, the Hoechst bright region h24
andthe N-banded region h25together havebeen
shown to contain the AAGAG and AAGAGAG sequences of the 1.705 satelliteDNA, the AAGAC
sequence of the 1.686 satellite and the AATAT sequence of the 1.672 satellite DNA (Figure 1). An
obvious suggestionis that region h24 is similar to the
other Hoechst bright regions and contains the AAT A T sequence, while region h25 contains the AAGAG, AAGAGAGand AAGAC repeats like the other
N-banded regions. The situation of region hlO-h13
with respect to the localization of the AAGAGsequence is much less clear. A likely possibility is that
the AAGAG sequence is contained only in region h10
and h12. However, we cannot exclude that these
repeats are located, perhaps in small amounts, also in
regions h l l and h13.
A puzzling problem is represented by region h4h6, a brightly fluorescent block which accommodates
a smallN-band (h5) and contains five repeated sequences belonging to three different satellite DNAs
(see Figure 1). Three of the five sequences havebeen
mapped throughout region h4-h6 since the lackof
breakpoints that further subdivide this interval made
it quite difficult to resolve the sequence distribution
within these regions (the only breakpoint available is
that of T l 3 , which, however, carries an additional
deficiency of region h5-h6). These three sequences
are the AAGAG and AAGAC sequences typical of N
bands and the AATAT sequence usually located in
the Hoechst bright regions. Thus it appears likely that
the first two sequences map to the N-banded region
h5 while theAATAT repeats are locatedin the
187
Hoechst bright regions h4 and h6. The two minor
1.672 repeats (AATAG and AATAC) that exclusively
map to these regions appear to be located on the two
sides of the breakpoint of T 1 3 , in region h4 and h6,
respectively (see Figure 1). However, due to the deficiency carried by T13, we cannot exclude that some
of these sequences are also locatedsomewhere within
region h5. The presence of AATAC repeats exclusively in region h5-h6 is rather surprising, since quantitations of these repeats showed that they comprise
nearly 10% of the Y (A. R. LOHE,A. J. HILLIKER
and
P. A. ROBERTS, unpublishedresults).Cytologically,
the entire region h4-h6 spans less then 10% of the Y
chromosome and nevertheless it contains four different satellitesequences in addition to the AATAC
repeats. The possibility exists therefore that the degree ofcompactionof the DNA contained in this
particular region is higher than in other regions of
the Y chromosome.
No satellite sequences have been localized in
the
Hoechst bright regions h17 and h 18,in regions h l 9
and h20 and in the N-banded region h14. Region
h20, which corresponds to the nucleolus organizer
constriction, contains the ribosomalDNAcistrons
(BONACCORSI,
PIMPINELLI
and GATTIunpublished).
Satellite sequences which have
not yet been identified
could be contained in the remaining regions. Alternatively, they couldbe organized differently from the
rest of the Y, accommodating long stretches of nonrepetitive or middle repetitive DNA.
Together theresults reported above clearlyindicate
that there is a good correlation between N-bands and
1.705 repeats and between brightly fluorescent regions and 1.672 sequences.However, heterochromatic blockscontaining different combinations of satellite repeats exhibit identical banding patterns. Thus
the molecular basis for banding does not appear to
depend solely on DNA content; otherfactors, like the
presence of specific proteins which can interact with
various repetitive DNAs in similar ways, might play a
major role in determining the response to banding
techniques.
Satellite DNA and Y chromosome fertility factors:
The Y chromosome of D. melanogaster carries six
complementation groups required for male fertility
(KENNISON1981; HAZELRIGG
et al. 1982; GATTIand
PIMPINELLI
1983). Four fertility factors havebeen
mapped to the long arm (kl-5, kl-3, kl-2and k l - I ) and
two to the short arm (ks-I and ks-2). Moreover, the
kl-5,kl-3 and ks-I fertility factors wereshown to
possess extremely large physicaldimensions. These
three loci are defined by a series of noncomplementing breakpoints and deficiencies distributed over
chromosomal regions containing up to 4000 kbof
DNA(GATTI and PIMPINELLI
1983; BONACCORSIet
al. 1988). The availablecytogenetic data permitted
the mapping ofk1-I, kl-2and b - 2 but not an estimation
188
S. Bonaccorsi and A. Lohe
of their size. The localization of the fertility factors,
different, fully fertile stocks (HALFER1981). Thus,
as results from the extensive analysis performed by
region h4-h6 could be a site of accumulation
of closely
GATTI and PIMPINELLI
(1983), is reportedonthe
related repetitive DNAs, which, at least in some popbottom of Figure 1.
ulations, is dispensable.
More recently it has been shown
that primary sperThe regions that correspond to the non-loop-formmatocyte nuclei of X/Y males of D. melanogaster exing geneskl-2 and ks-2 also contain satellite sequences.
hibit three giant lampbrush-like loops formed by the
ks-2 co-maps with 4 different repeats (AAGAG, AAY chromosome (BONACCORSI
et al. 1988). These strucGAGAG, AAGAC and AATAT), while kl-2 co-maps
tures are absent from X/O males and consist of three
with the AATAGAC, AAGAG and AAGAC repeats.
conspicuous skeins thread-like
of
material which begin
No evidencefor asubstantial presence of any of these
to develop in young spermatocytes, grow throughout
satellite sequences within kl-1 (region h14) has been
spermatocyte development and reach their maximum
obtained in the present analysis.In general, no specific
size in mature spermatocytes; they disintegrate just
association between a satellite sequence and a given
before the first meiotic division. Deficiency
and breakfertility factor can be observed. The same sequence
point mapping studies showed that the loop-forming
often maps to different fertility factors and each fersites map withinthe kl-5, kl-3and ks-1 fertility factors.
tility region is characterized by a specific combination
In particular, region h3, which spans about one third
of satellite sequencesrather than by an homogeneous
of kl-5 is responsible for the formation of the kl-5
array of a single type of repeats. Moreover, both the
loop, while region h2 1, which comprises one third of
loop-forming and the non-loop-forming segments of
ks-1, forms the ks-1 loop (Figure 1). The loop-forming
the kl-5 and ks-1 factors contain large amounts of
site of the kl-3 fertility factor is coextensive withregion
satellite sequences.
h7-h9 which corresponds to the minimum physical
It is currently very difficultto propose an hypothesis
et al.
sizeofthislocus
(Figure 1 and BONACCORSI
for the biological role of the satellite DNA contained
in the non-loop-forming fertility factors and in the
1988).
A comparison of the map position of the fertility
non-loop-forming regions of kl-5 and ks-1. Less diffifactors with the molecular map ofthe Y chromosome
cult is to envisage a possible function for the satellite
repeats contained in the loop-forming regions. The Y
constructed in the present workclearlyshows that
chromosome loops are giant nuclear structures conthese loci correspond to large blocks of satellite resisting of a DNA axis with laterally attached growing
peats(see Figure 1). The kl-5 locus contains four
transcripts in turn associated with large amounts of
different satellite DNA sequences; the loop-forming
proteins encoded by the autosomes and/orthe X
site of kl-5 (region h3) accomodates the AAGAG,
chromosome (BONACCORSI
et al. 1988; for the strucAAGAC and AAGAGAG sequences, while the kl-5
ture of the Y loops of D. hydei see HENNIG1985 and
non-loop-forming region h 1-h2 contains AATAT reLIFSCHYTZ
1988). I n situ hybridization experiments
peats. Similarly, four satellite sequences are located
on testes preparations have recently shown that the
within the ks-1 locus. The AAGAG and AAGAC
1.686-AAGAC sequence is actively transcribed along
sequences map to the loop-forming region h2 1; the
et al. 1990). The
the kl-5 and ks-1 loops (BONACCORSI
ks-1 non-loop-forming regions h22 and h23 contain
transcripts are accumulated on the loops and do not
the AATAAAC and AAGAG repeats, respectively.
appear to migrate to thecytoplasm; theyare degraded
Only AATAT repeats have been mapped to the klwhen the loops disintegrate prior to meioticmeta3 loop-forming region h7-h9, which corresponds to
phase
I. Togetherthese observations ledus to suggest
the minimum physical size of this fertility factor. Acthe
hypothesis
that these structures play a structural
tually, the physical size of kl-3 could exceed region
rather
than
a
coding role during spermatogenesis,
h7-h9 and possibly include part of region h4-h6.
providing
the
structural
framework for the compartUnfortunately, there are not currently available
mentalized
accumulation
of non-Y-encodedproteins.
breakpoints within the h4-h6 interval that permit a
better definition of the distal limit of this locus (GATTI The recent finding that thekl-3 loop binds a tektinS. BONACCORSI
and M. GATTI,
like protein (C. PISANO,
and PIMPINELLI
1983; BONACCORSI
et al. 1988). Thus
manuscript
in
preparation)
gives
further
support to
the kl-3 fertility factor might also contain a non-loopthis
hypothesis
and
indicates
that
some
of
the
proteins
forming portion corresponding to a region (h4-h6)
bound
to
the
loops
are
sperm-specific
polypeptides
to
which appears to contain at least fivedifferent satellite
be
utilized
later
in
development
during
sperm
differsequences (see above). In this respect it is interesting
entiation. An alternative hypothesis about the nature
to note that region h4-h6seems to be the only Y
of
the Y chromosome fertility factors has been put
chromosome block exhibiting a certain degree of cyforward
by GOLDSTEIN,
HARDY
and LINDSLEY
(1982)
tological polymorphism. Ananalysis carried out on
kl-5
and
kl-3
genes
who
observed
that
mutations
in
the
mitotic chromosomes of 16 stocks of D. melanogaster
lead
to
the
simultaneous
absence
of
two
different
high
demonstrated the presence of partial or complete
(M,
300,000)
and
of
molecular
weight
polypeptides
deletions of these regions of the Y chromosome in two
Satellite
Y Chromosome
the outer dynein arms of the peripheral doublets of
the axoneme. They interpreted these results as indicating that the kZ-? and kl-5 fertility genes contain the
coding sequences for the axonemal dyneins of Drosophila (GOLDSTEIN,
HARDYand LINDSLEY
1982). In
this respect, we would liketo point out thatour results
do not exclude the possibility that the kt-3 and kE-5
polypeptides observed by GOLDSTEIN,HARDYand
LINDSLEY
are encoded either by a segment of the loop
or by a region just outside the loop. Similarly, their
results do not exclude the possibility that, in addition
to coding for dyneins, the Y loops fulfill a protein
binding function.
We wish to thank M. GATTIand P. HARTE forhelpful discussion
and critical reading of the manuscript. This work has been s u p
ported in part by a grant from Fondazione Cenci Bolognetti, and
also by the Commonwealth Scientific and Industrial Research Organization, Australia.
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S., M. GATTI,
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C.
and A. LOHE,
1990 Transcription of a satellite DNA on two Y chromosome
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