J. CellSci. 7, 55-63 (1970)
Printed in Great Britain
55
THE FINE STRUCTURE OF MEIOTIC
CHROMOSOME PAIRING IN NATURAL AND
ARTIFICIAL LILIUM POLYPLOIDS
P. B. MOENS
York University, Downsvieiv, Ontario, Canada
SUMMARY
In the autotetraploid Lilmm longiflonim (411 = 48), there are 12 sets of 4 homologous chromosomes. Within each set of 4 homologues, switches of pairing partners and crossovers occur at
meiotic prophase. At the fine-structural level, the behaviour of the chromosomes is reflected in
the switches between the axial cores of the homologous chromosomes. The normal synaptonemal complexes of the autotetraploid are compared with the complexes of the allotriploid
L. tigrtnuvi, which have synaptonemal complexes with abnormal lateral elements. The possibility
that the deformed lateral elements are the products of heteromorphisms between' homologues'
is explored in the discussion. The observations on the chromosome cores are interpreted as
support for the notion that the cores may be associated with the recombinationally active
hereditary material during meiotic prophase.
INTRODUCTION
Paired homologous chromosomes normally have a synaptonemal complex during
the pachytene stage of meiotic prophase (Moses, 1968). Moses (1958) and Coleman &
Moses (1964) have suggested that the complex functions in the pairing of homologous
chromosomes and in crossing-over and chiasma formation. These functions are supported by Meyer's observations (1961, 1964) showing that where no genetic exchange
occurs, as in achiasmatic dipteran males, and in C3G D. melanogaster females, no
synaptonemal complexes occur. Callan (1967) proposes that the 4 master gene sequences
(Whitehouse, 1967) meet and undergo exchange inside the complex and the slave
sequences remain outside the complex, free from recombination. I used the same kind
of model to diagram the structure and function of the chromosomal cores in the
diploid Lihum longiflorum, at the leptotene, zygotene and pachytene stages of meiotic
prophase (Moens, 1968 a). In such models the axial core of the unpaired leptotene
chromosome and later the lateral element of the complex, because of its association
with the master gene sequence, depicts the orientation and behaviour of the recombinationally active genetic material during meiotic prophase.
The central role of the axial core is investigated here through a study of the chromosome pairing behaviour in the naturally occurring allotriploid Lilium tigrinum with
3 sets of chromosomes known to be less than fully homologous (Noda, 1966), and the
artificial autotetraploid L. longiflorum with 4 sets of homologous chromosomes. The
observations reported here support the view that the axial core represents the chromosome and possibly the recombinationally active genetic material in the various
associations.
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P.B.
Moens
MATERIALS AND METHODS
Artificially induced autotetraploids from diploid Easter lily, Ltltum longiflorum, were obtained
from De Graaf Bulbs, Gresham, Oregon. L. ttgrtnum (tiger lily) bulbs were obtained from a
local supplier. Anthers were fixed in 2 % glutaraldehyde solution in phosphate buffer for 2 h.
After a wash in buffer, the tissue was post-fixed in buffered 2 % osmium tetroxide solution for
1 h. Following dehydration through an alcohol series and propylene oxide, the anthers were
embedded in Epon 812, Luft's 1:1 mixture (Luft, 1961). Sections of about 60 ran thickness
were cut on an M T II Porter Blum microtome with a Dupont diamond knife. Serial sections
were collected according to Galey & Nilsson (1966), and deposited on Formvar-covered, single
hole grids. The sections were stained in a saturated aqueous solution of uranyl acetate and, after
washing, in lead hydroxide (Millonig, 1961). The sections were carbon coated and examined
with a Philips EM 200 at 60 kV. The 3-dimensional arrangement of chromosome cores and
complexes was determined by copying their positions in consecutive photographs on to a single
sheet of polyester drafting film.
OBSERVATIONS
The 4 homologous chromosomes of the autotetraploid lily can pair two by two, in
which case 2 bivalents are formed, each with a synaptonemal complex. Such complexes are normal in every respect and they are indistinguishable from those reported
for the diploid lily (Moens, 1968 a). In the triploid L. tigrinum. unusual lateral elements
can be seen in nearly every section of a pachytene nucleus (Fig. 2). The large number
of sections examined in the tetraploid showed no abnormal lateral elements. A change
of pairing partners in the autotetraploid is shown in Fig. 1. In the upper half of the
drawing (Fig. 1 A) the cores of homologues I and II form a synaptonemal complex as
do the cores of homologues II and IV. Below the switch homologues II and III are
paired, and homologues I and IV form a complex. The drawing summarizes the
segments of cores and complexes found in 12 consecutive sections. The numbers refer
to the sections in which the segments occurred. Sections 11, 10 and 8 are shown in
Figs. 1B, c and D respectively. Three other partner switches were traced and they
were similar to the one recorded in Fig. 1.
In the triploid lily, the cores of two of the homologues are paired into a synaptonemal
complex while the third, unpaired core follows along at some distance. Switches of
pairing partners occur regularly and have been described previously (Moens, 1969a).
The synaptonemal complexes of the triploid are unlike other complexes in that the
lateral elements frequently are branched or tubular (Fig. 2). The tubular structure was
confirmed in serial cross-sections of the complex (Fig. 2D). Additional characteristics
of the unusual lateral elements have been reported elsewhere (Moens, 19686) and are
referred to in the discussion.
DISCUSSION
If genetic exchange at meiosis is restricted to the confines of the synaptonemal
complex, then most of the chromatin is excluded from recombination by virtue of its
location on the outside of the complex. That some part of the genetic material may, in
fact, be excluded from recombination has been suggested by Callan (1967) on the
grounds that chiasmata do not involve the lateral loops of lampbrush chromosomes in
Chromosome pairing in Lilium polyploids
57
Triturus cristatus. Large interspecific variations in DNA content per genome (Rothfels
& Heimberger, 1968) imply that numerous genes are represented by several copies. In
the single-stranded model (Whitehouse, 1967) these copies are not detected genetically
(crossover or conversion) because they are excluded from recombination. Such
observations and models suggest that the genetic material of higher organisms at
meiotic prophase becomes partitioned into two distinct fractions, one participating in
recombination, and one excluded from recombination. I have speculated (Moens,
1968a) that the recombinationally active genes become organized on, or associated
with, the axial cores during leptotene and that the parallel alignment of the cores into
the lateral elements of the synaptonemal complex satisfies some of the physicochemical demands for genetic exchange to take place. If such an association between
the master gene sequence and the axial core exists, then the behaviour and the approximate location of master genes is reflected by the behaviour and position of the cores
during meiotic prophase. Thus the cores are expected to represent the chromosomes
in the usual, as well as the unusual, synaptic associations formed at meiotic prophase.
The behaviour of axial cores in normal meiotic prophase associations was demonstrated in the spermatocytes of Locusta migratoria (Moens, 1969c). Each of the
leptotene chromosomes has a continuous axial core attached with both ends to the
nuclear membrane at the polar region. The two cores of a pair of homologous chromosomes meet during zygotene to form a synaptonemal complex which is continuous
through the entire length of the bivalent and which is attached with both ends to the
nuclear membrane.
That the cores also represent the chromosomes in unusual situations was demonstrated in the triploid L. tigrinum (Moens, 1969a) where a change of pairing partners
between a set of 3 homologous chromosomes, as is commonly observed with the light
microscope, is mirrored at the fine-structural level by a change in axial cores. In an
autotetraploid, the 4 homologues can form multivalents, such as a chain of 4 or a ring
of 4. These configurations are the result of switches in pairing partners and crossovers
within the paired segments. It is shown in this report that the 4 cores of a set of 4 homologous chromosomes similarly switch pairing partners (Fig. 1). To one side of the
switch, cores I and II are paired as well as cores III and IV. After the switch of partners,
cores II and III form a synaptonemal complex, as do cores I and IV. In these cases
the axial cores reflect the essential cytological characteristics of the genetic material
expected from genetic and cytological observations in diploids and polyploids.
The remarkable difference in synaptonemal complexes of the triploid with deformed
lateral elements, and the tetraploid with normal elements, may be explained within
this framework. L. tigrinum is an allotriploid, derived from hybridization of two or
more closely related species (Stewart & Bamford, 1943; Noda, 1966). Unlike the
autotetraploid L. longiflorum, the allotriploid L. tigrinum has non-homologous segments
within a set of three homologues. Under the assumption that the axial core represents
the genetic material in meiotic associations, it is expected that inequalities in the
genetic material will be expressed as inequalities in the cores. Inequalities such as
short duplications or deletions would become visible only after synapsis takes place.
This interpretation agrees with the observation that in L. tigrinum the cores of the
58
P.B. Moens
unpaired leptotene chromosomes and the pachytene univalents are without deformities.
After two cores become the lateral elements of a synaptonemal complex the unusual
formations become apparent. In the autotetraploid the homologues are truly homologous and pairing does not produce abnormal lateral elements.
The speculation that the recombinationally active genes are associated with the
axial cores of meiotic prophase chromosomes is based on the general observation that
cores represent the chromosome in various synaptic relationships and one specific
observation, that heteromorphic' homologous' chromosomes pair and produce synaptonemal complexes with deformed lateral elements. The speculation can be tested
further by observations on the synaptonemal complexes of organisms with known
deletions, duplications, or other chromosomal mutations.
A number of observations bear witness against the model argued for here. Firstly,
the intergeneric hybrid Lycopersicon esculentum x Solanum lycopersicoides produces
few chiasmata, even though the chromosomes pair and form normal synaptonemal
complexes. The chromosomes of the haploid tomato exhibit a limited amount of
pairing with normal synaptonemal complexes (Menzel & Price, 1966). In these two
cases the obvious extensive non-homologies are not expressed as alterations of the
lateral elements. Secondly, one important feature of the master gene sequence is that
it undergoes recombination. There has, so far, been no report of a crossover or exchange between the lateral elements of the complex. Thirdly, if the recombinationally
active genes are arranged in a single linear sequence on the cores, their total length
would be only 100 to 1000 /im, depending on the organisms. This is only a minute
fraction of the total DNA per cell of a higher organism. Finally, core-like structures
and multiple cores are known to occur at times other than meiotic prophase and in
locations other than pachytene chromosomes (Roth, 1966; Wolstenholme & Meyer,
1966; Moens, 19696). Such cores are clearly not associated with the permanent genetic
material of the cell.
REFERENCES
CALLAN, H. G. (1967). The organization of genetic units in chromosomes. J Cell Sci. 2, 1-7.
COLEMAN, I. R. & MOSES, M J. (1964). DNA and the fine structure of synaptic chromosomes in
the domestic rooster (Gallus domesticus). J. Cell Biol. 23, 63-78.
GALEY, F. R. & NILSSON, C. E. G. (1966). A new method for transferring sections from the
liquid surface of the trough through staining solutions to the supporting film of a grid.
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LUFT, J. H (1961). Improvements in epoxy resin embedding methods. J. btophys. biochem.
Cytol. 9, 409.
MENZEL, M. Y. & PRICE, J. M. (1966). Fine structure of synapsed chromosomes in F1 Lycopersicon esculentum—Solatium lycopersicoides and its parents. Am. J. Bot. 53, 1079—1086.
MEYER, G. F. (1961). The fine structure of spermatocyte nuclei of Drosophtla melanogaster.
Proc. Europ. Reg. Conf. Electron Microsc, Vol. 2, pp. 951-954.
MEYER, G. F. (1964) A possible correlation between the submicroscopic structure of meiotic
chromosomes and crossing over. Electron Microscopy 1964 (ed. M. Titlbach), vol. B, pp. 4 6 1 462.
MILLONIG, G. (1961). A modified procedure for lead staining of thin sections. J. btophys.
biochem. Cytol. 11, 736-739.
MOENS, P. B. (1968a). The structure and function of the synaptinemal complex in Lilium
longiflorum sporocytes. Chromosoma 23, 418-451.
Chromosome pairing in Lilium polyploids
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MOENS, P. B. (19686). Synaptinemal complexes of Lilium tignnum (tnploid) sporocytes. Can.J.
Genet. Cytol. io, 799-807.
MOENS, P. B. (1969a). The fine structure of meiotic chromosome pairing in the tnploid,
Lihum tigrinum. J. Cell Biol. 40, 273-279.
MOENS, P. B. (19696). Multiple core complexes in grasshopper spermatocytes and spermatids.
J. Cell Biol. 40, 542-551.
MOENS, P. B. (1969c). The fine structure of meiotic chromosome polarization and pairing in
Loctista rmgratona spermatocytes. Chromosoma 28, 1—25.
MOSES, M. J. (1958). The relation between the axial complex of meiotic prophase chromosomes
and chromosome pairing in a salamander {Plethodon cinereus). J. biophys. biochem. Cytol. 4,
633-638.
MOSES, M. J. (1968). Synaptinemal complex. A. Rev. Genet. (Annual reviews, Palo Alto) 2,
363-412.
NODA, S. (1966). Cytogenetics on the origin of triploid Ltlium tignnum. Bull. Osaka Gaktun
Univ. 6, 85-140.
ROTH, T . F. (1966). Changes in the synaptinemal complex during meiotic prophase in mosquito
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ROTHFELS, K. & HEIMBERGER, M. (1968). Chromosome size and DNA values in Sundews
(Droseraceae). Chromosoma 25, 96-103.
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(Received 24 October 1969)
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P. B. Moens
Fig. i. A switch between the 4 painng partners in the sporocyte of the autotetraploid
Lihum longiflorum is mirrored at the fine-structural level by a switch between the
chromosomal cores, A is a diagram summarizing the segments of synaptonemal complexes and axial cores encountered in 12 consecutive sections of a partner switch. The
arable numbers refer to the sections in which the segments occur. The roman numbers
identify the cores of the 4 homologues. The core of homologue I first forms a synaptonemal complex with the core of homologue I I ; it then switches over and forms a complex with the core of homologue IV. Core III similarly switches from IV to II. Where
the cores are paired into a synaptonemal complex the central element ce is present.
B, c and D are electron micrographs of sections 11, 10 and 8 respectively. The scale
line on Fig. I B represents 0-5 /im and Figs. 1 c, D are at the same magnification.
Chromosome pairing in Lilium polyploids
.tc
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P. B. Moens
Fig. 2. Synaptonemal complexes in the sporocyte of the allotnploid Lilium tignnum.
Complexes are formed between the cores of 2 of a set of 3 homologues and they have
abnormal lateral elements, A, A synaptonemal complex with a normal lateral element
(le) and a deformed lateral element (die). The chromatin (cli) of this cell in the zygotene
stage of meiotic prophase is still loosely arranged around the complex with central
element (ce). Not shown is the third homologue with a single core which follows along
the bivalent at some distance from it. B, A typical deformed lateral element (die) in a
mid-pachytene cell with condensed chromatin (cli). c, A cross-section through a synaptonemal complex with a normal lateral element (le), a central element (ce) and a
deformed lateral element (die), D, One electron micrograph of a series of consecutive
sections, showing that this deformed lateral element is a hollow spindle.
Chromosome pairing in Lilium polyploids