The mouse salivary androgen-binding protein (ABP) gene
cluster on Chromosomes 7: characterization and
evolutionary relationships
Christina M. Laukaitis,1,* Stephen R. Dlouhy,2 Robert C. Karn1
1
Department of Biological Sciences, Butler University, 4600 Sunset Avenue, Indianapolis, Indiana 46208, USA
Department of Medical and Molecular Genetics, Indiana University School of Medicine, 975 W. Walnut, Room 130,
Indianapolis, Indiana 46202, USA
2
Received: 18 April 2003 / Accepted: 18 June 2003
Abstract
Mouse salivary androgen-binding protein (ABP) is a
pair of dimers, composed of an alpha subunit disulfide bridged to either a beta or a gamma subunit. It
has been proposed that each subunit is encoded by a
distinct gene: Abpa, Abpb, and Abpg for the alpha,
beta, and gamma subunits, respectively. We report
here the structures and sequences of the genes that
encode these three subunits. Each gene has three
exons separated by two introns. Mouse salivary ABP
is a member of the secretoglobin family, and we
compare the structure of the three ABP subunit
genes to those of 18 other mammalian secretoglobins. We map the three genes as a gene cluster located 10 cM from the centromere of Chromosome
(Chr) 7 and show that Abpa is the closest of the three
to the gene for glucose phosphate isomerase (GPI)
and that Abpg is the closest to the centromere, with
Abpb mapping between them. Abpa is oriented in
the opposite direction to Abpb and Abpg, with its 5¢
end directed toward their 5¢ ends. We compare the
location of these genes with other secretoglobin
genes in the mouse genome and with the known
locations of secretoglobin genes in the human genome and present evidence that strong positive selection has driven the divergence of the coding
regions of Abpb and Abpg since the putative duplication event that created them.
The nucleotide sequence data reported in this paper have been
submitted to GenBank and have been assigned the accession
numbers Abpa: AF144714; Abpb: AY325897; Abpg: 325898.
*Current address: Department of Internal Medicine, St. Vincent
Hospital, 2100 W. 86th St., Indianapolis, IN 46260, USA
Correspondence to: R.C. Karn; E-mail: [email protected]
A gene’s location within the genome reflects its evolutionary history and affects its expression. With
the recent genome sequencing effort, it has become
possible to study the context surrounding individual
genes. In a simplistic model, gene duplications
would initially place genes within close proximity to
one another within the genome where they potentially could be coordinately up- or down-regulated.
Indeed, in the zebrafish, the cluster of evolutionarily
related odorant receptor genes are found in close
proximity, and individual chromosomal clusters are
expressed in the same olfactory epithelial zone
(Kratz et al. 2002), suggesting both gene duplication
and coordinate regulation. More direct evidence for
coordinate regulation comes from the occurrence of
chromosome remodeling of an entire cluster of genes
before expression of those genes. For example, the
genes for cytokines IL-4, IL-5, and IL-13, which are
required for TH2 differentiation, are located in a 180kb region of human Chr 5, and this region of chromatin undergoes remodeling before TH2 differentiation (Agarwal et al. 1999).
We are interested in the murine members of the
secretoglobin protein family (Klug et al., 2000) that
are expressed in salivary glands. This family, called
mouse salivary androgen-binding protein (ABP),
consists of two different dimers: an alpha subunit is
bound to either a beta or a gamma subunit by disulfide bridging before secretion into mouse saliva
(Dlouhy et al. 1987). From biochemical genetic evidence, Dlouhy et al. (1987) postulated that three
distinct genes, Abpa, Abpb, and Abpg, encoded the
three protein subunits, alpha, beta, and gamma, respectively. They obtained three products with distinctly different isoelectric points from cell-free
translation of a submaxillary gland poly(A)-tailed
RNA extract, suggesting that each of the three
DOI: 10.1007/s00335-003-2291-y • Volume 14, 679–691 (2003) • Springer-Verlag New York, Inc. 2003
679
680
C.M. L AUKAITIS
subunits was translated from a different mRNA and
lending additional support to the hypothesis of three
ABP genes. They could not exclude, however, the
possibility that exon shuffling was responsible for
the different mRNAs.
More recently, the cDNAs for these subunits
have been described (Karn and Russell 1993; Karn
and Laukaitis 2003), and the amino acid sequences
predicted from them suggest that these three proteins are evolutionarily related to each other and to
other secretoglobins. Construction of a phylogenetic
tree suggests that Abpa diverged from a common
ancestor of Abpb and Abpg, and thus Karn and
Laukaitis (2003) proposed that the three putative
genes originated from gene duplication events. Previous genetic studies have suggested that Abpa and
Abpg are closely linked and assigned Abpa to Chr 7,
at a location 10 cM from the centromere (Dlouhy
et al. 1987), but the lack of electrophoretic polymorphism of Abpb precluded mapping it with
standard recombination techniques. If the hypothesis of gene duplication is correct, then Abpa, Abpb,
and Abpg might still be found in close proximity to
one another within the genome.
It was our purpose in this study to resolve the
issue of the number of genes encoding mouse salivary ABP subunits and to produce a physical map of
the gene structures and the organization of the
cluster. We demonstrate the existence of three distinct genes encoding the alpha, beta, and gamma
subunit cDNAs and show that these are clustered on
Chr 7 in the mouse. We also characterize the exon
and intron structures and orientations of these three
genes. There is evidence that one or more of these
genes have evolved under strong Darwinian selection (Hwang et al. 1997; Karn and Nachman 1999;
Karn et al. 2002), and we examine the potential evolutionary relationships among these genes and
other secretoglobin genes.
(Sigma Biochemicals, St. Louis, Mo.) as previously
described (Hwang et al. 1997; Karn and Nachman
1999). Long-range PCR was performed with the LA
PCR kit from TaKaRa Biotechnical Supplies (Osaka,
Japan), using recipes recommended by the manufacturer. The reaction included: 1 min at 94C; 14
cycles of 20 s at 98C, 20 s at 50C, and 10 min at
68C; 6 cycles of 20 s at 98C, 20 s at 56C, and 11
min at 68C; 6 cycles of 20 s at 98C, 20 s at 56C,
and 12 min 30 s at 68C; 6 cycles of 20 s at 98C; 20 s
at 56C, and 14 min at 68C; and 5 min at 72C.
Materials and methods
Polymerase chain reaction. Templates for PCR reactions were genomic DNAs from the inbred DBA/2J
and C3H/HeJ mouse strains purchased from The
Jackson Laboratory (Bar Harbor, Maine). Production
of submaxillary gland cDNA libraries has been previously described (Hwang et al. 1997; Karn and
Laukaitis 2003). Primers for amplification of Abpa,
Abpb, and Abpg were designed from their cDNAs
(Karn and Laukaitis 2003) and purchased from Genosys (St. Louis, Mo.). Their positions are shown in
the gene figures, where they are underlined with the
direction of their 3¢ ends indicated with arrows. PCR
was performed by using Sigma Taq Polymerase
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
DNA sequencing. Clean-up of PCR templates
was performed by using QiaQuick PCR clean-up
spin columns according to the manufacturer’s protocol (Qiagen Incorporated, Valencia, Calif.). Automated sequencing was performed by ACGT, Inc.
(Chicago, Ill), and manual sequencing was performed
by using Amersham end-termination sequencing
kits (Amersham-Pharmacia, Uppsala, Sweden; Karn
and Nachman 1999).
Data analysis. DNASIS/PROSIS v. 5.0 (Hitachi
Software, Yokohama, Japan) was used to search and
to manually align DNA sequences isolated from
Blast searches as well as in primer design. The program MEGA, version 2.1 (Kumar et al. 2001), was
used to calculate nucleotide divergences with the
Kimura correction for multiple hits and a transition:
transversion ratio of 2. The distances and their
standard errors were compared by a modification of a
one-tailed t-test with infinite degrees of freedom
(Graur and Li 2000).
Genome mining. High throughput genome sequencing data from mice were searched by using Blast
(Altschul et al. 1990) for sequences corresponding to
cDNA sequences of Abpa, Abpb, and Abpg (Karn and
Laukaitis 2003). BACs identified as having scores
>200 were downloaded for further analysis. The Ensembl program (Hubbard et al. 2002) was used with
the February 2002 mouse genome assembly to visualize the assigned locations of DNA contigs, unsequenced gaps, known genes, and gene predictions
on the proximal end of Chr 7. The synteny-view feature was used to assess homology between secretoglobin-containing regions of mouse Chrs 7, 18, and
19 and human Chrs 5, 11, and 19. BLAT (Kent 2002)
was used to search the February 2002 mouse genome
assembly to determine the chromosomal location of
Abpg. The UCSC genome browser (Karolchik et al.
2003) was used to search for CpG islands, GC content, gene predictions, and sequence repeats in proximal Chr 7 and to obtain DNA sequence of the 80 kb
region spanning Abpg through Abpa.
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
681
C HR 7
Table 1. Exon and intron structures for 21 secretoglobin chains
Exon 1
Group
A
B
C
D
E
Gene
Total Size
(kb)
Oc Utero
Hs CC10
Mm CC10
Rn CC10
Mm Abpa
Mm lachrymal
Fc Fel dI, chain 1
Hs MGB A
Hs MGB B
Rn PBP C3
Ma HGB B1
Ma HGB B2
Ma HGB A
Rn PBP C1
Rn PBP C2
Hs LPP A
Hs LPP B
Fc Fel dI, chain 2
Mm 11 kDa
Mm Abpb
Mm Abpg
5¢ UTR
Coding
Exon 3
Total Intron 1 Exon 2 Intron 2
3.08
47
55
102
2286
188
4.11
8
55
63
3070
188
4.33
72
45
127
2471
188
3.55
57
55
112
1680
188
1.33
20a
61
81
121
188
1.30
16
64
80
119
188
N/A
?
61
N/A
155
188
3.00
59
56
114
609
188
5.26
64
55
119
1612
188
2.76
54
58
112
1551
188
Only cDNA sequences are avalible at this time
Only cDNA sequences are avalible at this time
Only cDNA sequences are avalible at this time
3.01
36
61
97
1719
191
2.88
33
52
85
1634
203
3.25
47
55
102
1716
188
2.55
54
55
109
926
188
N/A
?
61
N/A
512
182
N/A
?
61
N/A
465
182
1.99
31a
61
92
451
182
1.99
31a
61
92
451
182
Coding
3¢ UTR
Total
30
161
45
45
27
27
27
35
42
39
143
254
127
118
128
123
(Term)
164
169
170
173
137
172
163
155
150
N/A
199
211
209
859
81
805
81
1105
27
1193
27
1251
78
Not determined
993
93
1009
93
61
68
112
111
(Term)
142
149
139
138
N/A
161
161
254
254
327
650
1376
1406
785
761
1101
1888
3128
695
a
Based on putative 5¢ end.
BLAT was used to obtain gene sequences for other
secretoglobins in mouse, rat, and human. cDNA sequences were searched against the November 2002
human, November 2002 rat, or February 2002 mouse
genome assemblies, as appropriate, to determine exon
and intron size and placement. 5¢ and 3¢ UTR assignments were performed according to ATG and stop codon locations in the gene and polyadenlyation signal
sites in the cDNA. GenBank cDNA accession numbers are: mouse ABP alpha (AF144714), mouse lachrymal (AF008595), rat PBP C1 (V01255), rat PBP C2
(V01256), rat PBP C3 (V01263), human lipophilin A
(AJ224171), human lipophilin B (AJ224172), human
mammoglobin A (U33147), human mammoglobin B
(AF071219), human Clara 10 (X13197), mouse Clara 10
(L04503), rat Clara 10 (NM_013051), hamster heteroglobin A (Z66540), hamster heteroblogin B1
(AJ252138), and hamster heteroglobin B2 (AJ252139).
No gene corresponding to an exact match for the mouse
11-kDa cDNA (AF272844) was found; gene structure
was predicted in Table 1 from a similar predicted gene
with 90% homology (ENMUSG00000036047). The
rabbit uteroglobin gene structure (X01423) was previously determined, as were partial gene structures for
feline Fel d I chain 1 (X62477) and chain 2 (X62478).
Results
The structures of the ABP genes (Abpa, Abpb and
Abpg). The three genes encoding the subunits of
mouse salivary ABP are known to be expressed in
mouse submaxillary glands since mRNA encoding
all three subunits can be isolated from the glands
(Dlouhy and Karn 1984; Hwang et al. 1997; Karn and
Laukaitis 2003), and protein composed of the alpha
subunit bound to either the beta or gamma subunits
can be isolated from submaxillary secretions (Karn
and Russell 1993; Karn and Clements 1999). Abpa,
the gene common to all forms of ABP described to
date, has been until now only partially sequenced.
Karn and Russell (1993) reported the sequence of a
partial cDNA that included the coding region of the
entire secreted alpha subunit and the 3¢ UTR. The
intron/exon structure of Abpa was delineated and
the sequence of the second intron produced by Karn
and Nachman (1999). We have completed the sequence of Abpa. Although the 5¢ UTR of the Abpa
cDNA was not known previously, we were able to
design a primer for a putative 5¢ UTR using sequence
in the mouse genome. Using this primer and a reverse primer in the second exon, we were able to get
amplification from a submaxillary cDNA template
(Fig. 1), and we confirmed that this represented expressed Abpa by sequence analysis of the coding
region representing the secreted protein (not shown).
We were also able to determine the signal peptide.
We subsequently utilized these primers for amplification from genomic DNA (Fig. 1) and determined
the sequence of the first intron of Abpa. Thus, our
data complete the structure of Abpa by adding the
682
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
Fig. 1. Abpa and Abpb amplification products. Panel A: Amplification of the gene corresponding to the complete cDNA of
Abpa. Lane 1, molecular weight standards; lane 2, a 1.0-kb portion of Abpa amplified from genomic DNA with a forward
primer in Exon 2 and a reverse primer in the 3¢ UTR (Karn and Nachman 1999); lane 3, the partial cDNA amplified using
the same primers as in lane 2; lane 4, the full-length (1.33 kb) Abpa amplifed from genomic DNA with a forward primer in
the 5¢ UTR, a reverse primer in the 3’ UTR; and lane 5, the full-length cDNA amplified with the same primers as in lane 4.
Panel B: Sequencing strategy for the Abpb gene (all PCR performed on genomic DNA). Lane 1, molecular weight
standards; lane 2, the 0.6-kb 5¢ end of Abpb amplified with a forward primer in the 5’ UTR and a reverse primer in Exon 2;
lane 3, the 1.3-kb 3¢ end of Abpb amplified with a forward primer in Exon 2 and a reverse primer in the 3¢ UTR; and lane 4,
the ca. 2-kb doublet obtained when PCR is performed on genomic DNA by using a forward primer in the 5¢ UTR and a
reverse primer in the 3’ UTR. Panel C: Amplification of the fragment separating Abpa and Abpb. Lane 1, molecular weight
standards; lane 2, the ca. 8-kb fragment separating Abpa and Abpb, amplified from genomic DNA by using primers at the
5¢ ends of both genes. These were pointed toward the 5¢ ends of the two genes and are diagrammed on Fig. 5.
sequences of the first exon and the first intron
(Fig. 2). The first exon includes a small 5¢ UTR and
the coding sequence for most of the signal peptide,
and this is followed by a very small intron (121 bp).
The cDNA result shown in Fig. 1A, lane 5, confirms
that the additional exon has been correctly assigned.
We also report here the complete structures of the
beta subunit gene, Abpb (Fig. 3) and the gamma
subunit gene, Abpg (Fig. 4). We began with a strategy
similar to that of Karn and Russell (1993). Briefly,
mixed oligomer probes were designed by using a
partial beta subunit protein sequence (Karn and
Russell 1993) and were used to probe a submaxillary
cDNA library for complementary clones which were
sequenced. A cDNA, putatively identified as encoding the beta subunit of ABP including part of its
transcription initiation sequence, was obtained and
sequenced (Karn and Laukaitis 2003). Confirmation
of its identity required comparison with that encoding the alternative subunit, gamma, since the protein
sequences of these were reported to be quite similar
(Karn and Clements 1999). Primers designed from the
5¢ and 3¢ UTR sequences of the cDNA yielded a
doublet of bands of approximately 2 kb when used to
amplify genomic DNA (Fig. 1). To ensure that we
sequenced only a single gene corresponding to the
cDNA, we designed a set of antiparallel but partially
overlapping primers near the center of the coding
region (eventually found to be in Exon 2; Fig. 3) and
used each in conjunction with the appropriate 5¢ or 3¢
UTR primer to amplify fragments of the gene (Fig. 1).
These fragments were sequenced and assembled to
yield a gene approximately 2 kb in length (Fig. 3). The
subsequent release of the mouse genome allowed us
to locate this gene in the database.
Using a similar strategy to that outlined for
Abpb, we designed a mixed oligermeric DNA probe
to re-probe the cDNA library for clones encoding the
gamma subunit. We subsequently obtained a putative gamma cDNA sequence and showed that the 5¢
UTRs, signal peptide coding regions, and the 3¢
UTRs of the beta and gamma cDNAs are almost
identical (Karn and Laukaitis 2003). This similarity
explains why we observed two bands when amplifying genomic DNA with primers located in the 5¢
and 3¢ UTRs. Identifying the gene shown in Fig. 3 as
Abpb was possible when the ‘‘a’’ and ‘‘b’’ alleles of
the beta and gamma cDNAs were sequenced from
C3H/HeJ and DBA/2J mouse submaxillary glands,
respectively (Karn and Laukaitis 2003). Identification required comparing a protein sequence encoded
by both subunits in both strains to predict the electrophoretic variations upon which the original subunit assignment was made (Dlouhy et al. 1987). We
deduced the gamma gene (Abpg; Fig. 4) from the
mouse genome, using its cDNA sequence (Karn and
Laukaitis 2003). The exons were assigned from the
cDNA coding sequences and their 5¢ and 3¢ UTRs,
and the intron boundaries were verified by the consensus GT and AG doublets flanking the ends of the
exons.
The sequences of Abpa, Abpb, and Abpg, which
are given in Figs. 2–4, respectively, show that each is
composed of three exons and two introns, which are
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
683
Fig. 2. Abpa, the mouse androgen-binding protein alpha subunit gene. The gene consists of three exons and two introns
shown in the diagram above the sequence. The amino acid sequence encoded by the exons are written below them. The
putative primary promoter region, a potential secondary promoter region much farther upstream, and the polyadenylation
signal are boxed. Transcription initiation sequences are in bold. GT/AG splice sequences in the introns are in italics.
Primer sequences are underlined with arrows indicating their 3¢ ends.
very similar in size between Abpb and Abpg. By
contrast, these segments in Abpa have much different sizes and sequences:
1. The coding regions of Exon 1 are the same size
for all three subunits, but the sequence of Abpa is
different from the other two, which are nearly
identical to each other (92% with one insertion/
deletion [indel]). The predicted 5¢ UTR size varies by
11 bp between Abpa (20 bp) and Abpb and Abpg
(31 bp).
2. Intron 1 is much smaller for Abpa (121 bp)
than for the other two (451 bp), and it has essentially
no sequence in common with them. By contrast,
there is significant identity (90%) between Abpb and
Abpg in this intron.
3. Exon 2 is similar in size (188 bp in Abpa; 182
bp in Abpb and Abpg) among the three subunit
genes, but the sequence of Abpa has essentially no
sequence in common with Abpb and Abpg (which
are 72% similar to each other), as reported for their
cDNAs (Karn and Laukaitis 2003).
4. Intron 2 is smaller in Abpa (785 bp; [Karn and
Nachman 1999]) than in Abpb (993 bp) and Abpg
(1009 bp) and shows no significant sequence identity
with them. Intron 2 of Abpb and Abpg, on the other
hand, shows 90% identity; the size differences ac-
684
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
Fig. 3. Abpb the mouse androgen-binding protein beta subunit gene. The gene consists of three exons and two introns
shown in the diagram above the sequence. Control regions are boxed, and other labels are as described for Fig. 2.
counted for by 5 indels, the largest of which is 14 bp.
The net effect of the 5 indels distributed between the
Abpb and Abpg second introns is a size difference of
only 16 bases.
5. Exon 3 is smaller in Abpa (155 bp) than Exon 3
in Abpb and Abpg (254 bp in each). The region
contains the end of the coding sequence (27 bp in
Abpa and 93 bp in both Abpb and Abpg) and the 3¢
UTR (as determined from cDNA sequences; 146 bp
in Abpa, 170 bp in Abpb, and 171 bp in Abpg). Exon
3 of Abpa shows no significant identity with the
other two. Abpb and Abpg share 61% identity in the
Exon 3 portion of their coding regions and 89% in
their 3¢ UTRs.
Putative control regions of Abpa, Abpb and
Abpg. Figures 2–4 also show putative control regions for the three ABP subunit genes, which were
surmised 1) from the distances from the putative
promoter to the putative transcription initiation site
and 2) from consensus sequences derived from other
1 studies (White 2001; Klug and Cummings 2003).
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
685
C HR 7
Fig. 4. Abpg, the mouse androgen-binding protein gamma subunit gene. The gene consists of three exons and two introns
shown in the diagram above the sequence. Control regions are boxed and other labels are as described for Fig. 2.
Abpa has a sequence (TATAAAA) consistent with
the primary promoter consensus TATAAT 25 bp
proximal to the transcription initiation site (GAGCACTTGCC). It does not have a CAAT consensus
80 bp proximal to that site, the expected position of
the secondary promoter sequence, but it does have a
CAAT sequence 205 bp proximal to the transcription initiation site. Abpb and Abpg have identical
sequences (TGATAAT) consistent with the primary
promoter consensus 25 bp proximal to the transcription initiation site (GCTCTGAGAAGAGC in
Abpb and GCTTTGAGAAGAGC in Abpg). They
also both have a putative secondary promoter sequence (GCTCAAAGCT in Abpb and GCCCAA
AGGT in Abpg) 80 bp proximal to that site. Using a
UCSC genome browser search tool, we found no
CpG islands or GC-rich regions that are commonly
associated with housekeeping genes in the area immediately proximal to the transcription initiation
sites. These findings are consistent with the fact that
ABP genes are primarily expressed in salivary glands
(Dlouhy et al. 1986).
686
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
Fig. 5. The structure of the mouse salivary androgen-binding protein (ABP) gene cluster on Chr 7 with the contigs used to
construct the map shown below the chromosome bar. The centromere is to the left (24.5 Mb or ca. 10 cM) of the center of
the ABP cluster. The structures of Abpb and Abpa are shown above the chromosome bar, with their 5¢ ends oriented
towards each other. The primers used in long-range amplification are shown above the Abpb and Abpa gene structures.
The structure of the Abpg gene is shown below the chromosome bar, mapping in the same 5¢-to-3¢ orientation as Abpb and
to the left of Abpb and Abpa. The exons of each gene are shown with shaded coding regions and unshaded noncoding
regions (5¢ and 3¢ UTRs). The sizes of each gene are shown below its diagram. Also shown are the Mus musculus Major
allergen 1 (Mjal 1) gene on the left flank of the ABP cluster, the glucose phosphate isomerase (GPI) gene on the right flank
of the cluster, and five LINE 1 (L1) genes interspersed among the ABP genes.
The physical map of the ABP gene cluster. A
BLAST search of the Mus musculus high-throughput
genome sequencing database for Abpa identified
three BACs containing full-length matches with
greater than 99% homology: RP23-69B1, RP23291N20, and RP23-477F12. A similar BLAST search
for Abpb identified the same three BACs. A search
for Abpg identified only RP23-477F12. All of these
BACs have been mapped to the proximal end of
mouse Chr 7 by Ensembl. The individual Abpa and
Abpb gene sequences are located on separate subclones within RP23-291N20 and RP23-69B1 and on
the same clone within RP23-477F12. This places
Abpa and Abpb approximately 8 kb apart with their
5¢ regions oriented toward each other and their 3¢
ends pointing away from each other (Fig. 5). The
sequence predicted to fall between Abpa and Abpb
corresponds to the ends of the two subclones con-
taining Abpa and Abpb in RP23-291N20 and RP2369B1. We confirmed the orientation of Abpa and
Abpb relative to each other by amplifying an overlapping segment using long-range PCR with genomic
DNA as the template (Fig. 1). We obtained a PCR
product of the expected 8-kb size when we amplified
with primers pointed toward the 5¢ direction of both
Abpa and Abpb (Fig. 5), but with none of the other
possible primer combinations.
Abpg is found on a single clone within RP23477F12. This subclone contains neither Abpa
nor Abpb. The ends of this piece showed no
homology to any of the other subclones identified as
containing Abpa or Abpb. The Ensembl database
maps RP23-477F12 immediately proximal to RP2329N120. The appearance of Abpg in this region is
consistent with the earlier linkage of Abpa and Abpg
with recombination data (Dlouhy et al. 1987) and
Fig. 6. Comparison of nucleotide divergences for different segments of Abpb and Abpg. The diagram at the top of the figure
shows the general structure of the two genes. The nucleotide divergences with standard errors for the exons and introns
are shown below the diagram. Nucleotide divergences for regions within the three exons are shown at the bottom.
C HR 7
687
supports the location of this contig in this segment
of Chr 7.
Comparing the map we derived with the map of
this region assembled by Ensembl shows that Abpb
is closest to the centromere, while Abpa is closest to
GPI (Fig. 5). BLAT was used to search the UCSC
mouse database for Abpa, Abpb, and Abpg, and
confirmed the assignment of Abpa to nucleotides
24,507,099 through 24,508,667 of Chr 7, running in a
5¢ to 3¢ orientation. The Abpa sequence described in
the genome databases differs in the sequence of its
second intron by two bases from our sequence. These
are at positions 912 and 1126, and are emboldened
and underlined on Fig. 2. Current and previous DNA
sequencing (Karn and Nachman 1999) support our
version of Intron 2. It must be noted, however, that
we are working with the C3H/HeJ strain of mice
while genome sequencing used the C57BL/6 strain.
Both strains are Abpaa/Abpaa (Dlouhy and Karn
1983), but minor differences in their genes could exist. A BLAT search of the UCSC database retrieved a
sequence essentially identical to Abpb found on Chr
7 with its 3¢ end at 24,497,716 and its 5¢ end at
24,499,896. This distance confirms our placements,
including relative orientations, of Abpa and Abpb.
One difference exists between the genome sequences
at this location and the sequence we report here for
Abpb. This change is at bp 280 and is emboldened
and underlined. Here, we report a T for the genome
project’s C. The C at this position would change the
amino acid of the signal peptide, substituting a Thr
for an Ile, which is inconsistent with both our current
DNA sequencing data (Karn and Laukaitis 2003) and
with our previous protein sequence (Karn and Russell
1993; Karn and Clements 1999); however, all that
work with the ‘‘a’’ variant of ABP a has been done
with the C3H strain. A BLAT search of the UCSC
database retrieved a sequence 100% identical to the
cDNA of Abpg with its 3¢ end at 24,429,469 and its 5¢
2 end at 24,431,662 on Chr 7. This places the 5¢ end of
Abpg 66.054 kb from the 3¢ end of Abpb in the same
orientation (Fig. 5).
The gene prediction program Genescan embedded within the UCSC genome browser predicts the
existence of Abpb (labeled Chr7_9.8) and the first
two exons of Abpg, although the third exon is
incorrectly predicted (Chr7_9.6). Ensembl gene prediction software does not predict either Abpb or
Abpg. Thus, our current and previous work with
protein and cDNA sequences has enabled us to identify all three genes (Abpa, Abpb, and Abpg) that are
expressed in adult submaxillary glands. A number of
other features are found in close proximity to the
ABP genes on mouse Chr 7. At least four repeats of
4 kb in length flank the ABP gene cluster. These
repeats are 92–99% homologous to a retrotransposon
L1 (AF081104) and are plotted on Fig. 5. A number of
genes with 90% homology to Abpa have also been
found in this area (Waterston et al. 2002) and we have
noted numerous Abpb and Abpg paralogs.
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
Comparison of Abpa, Abpb and Abpg with
other secretoglobin genes. Karn and Laukaitis
(2003) compared the amino acid sequences of 21 secretoglobin chains from various mammalian species
and derived a molecular phylogeny consistent with
the five groups (A–E; see Table 1) of Ni et al. (2000),
adding the M. musculus lachrymal proteins and expanding group E to include the Mus musculus salivary ABP beta and gamma subunits. We identified
gene sequences encoding the 21 proteins clustered in
groups A–E by Karn and Laukaitis (2003) and compared their intron and exon sizes (Table 1). Overall,
the structures of the 21 secretoglobins we studied are
similar: three exons separated by two introns. The
sizes of these genes differ widely, however, from
about 2 kb to over 5 kb with a mean of 2.96 (S.D.
1.08). Most of the breadth of range is owed to the
highly variable introns. The region least variable in
size is in the middle exon (mean 188 bp, S.D. 4 bp).
Exons 1 and 3 show more size variation (Exon 1:
mean 99 bp, S.D. 17; Exon 3: mean 176 bp, S.D. 45),
owing mostly to variability in the sizes of the 5¢ and
3¢ UTRs, although we realize that variability in 5¢
and 3¢ UTRs may be due at least in part to how
complete these regions were in the reported cDNA
sequences. The introns vary much more widely in
size than the exons (Intron 1: mean 1197 bp, S.D.
892; Intron 2: mean 1137 bp, S.D. 1066).
Comparing the phylogeny developed on the basis
of amino acid sequence similarity of their products
to the exon/intron structures of the genes reveals
interesting relationships. Group A, the uteroglobin/
claras, has the largest average gene size (mean 3767
bp, S.D. 567). Group C comes close to this average
(mean 3670 bp, S.D. 1380) only because of the very
large second intron in Hs MGB B, making it the
largest gene of the 21. Otherwise, group C gene sizes
are closer to the group D average size (mean 2922 bp,
S.D. 290). Groups B (mean 1314 bp, S.D. 23) and E
(mean 1982 bp, S.D. 8), the ones containing the ABP
genes, are much smaller than the other three groups,
although 5¢ and 3¢ UTR sizes cannot presently be
determined for one member of group B and two of
group E. Perhaps the most interesting observation
about group E is that all of the genes have their
middle exon reduced by six nucleotides, three on the
5’ end and three on the 3¢ end. This coincides with a
shift in the cleavage point of the signal peptide such
that the first Cys in the amino acid sequence has
688
C.M. L AUKAITIS
become residue 1 in the secreted forms of group E,
while it is residue 3 in all the other sequences.
Laukaitis 2003). The divergences of the 5¢ and 3¢
UTRs are comparable to the intron values, as is that
for Helix 3. The other three helices have significantly higher values (P < 0.01; one-tailed t-test with
infinite degrees of freedom), suggesting that they
represent the regions of the secreted proteins that
have been under strong positive selection.
Nucleotide divergence of Abpb and Abpg. The
nearly identical structures of Abpb and Abpg suggest
that they are the products of gene duplication. Surprisingly, their first exons, which contain the coding
regions for nearly all of the signal peptides, and their
introns are much more similar than the coding regions (Exons 2 and 3) for their secreted protein products as shown by their nucleotide divergences (Fig. 6).
In our previous work, we showed that positive selection operated on the coding sequences of the secreted proteins by demonstrating relatively high
nonsynonymous/synonymous substitution rate ratios (Ka/Ks) in pairwise comparisons of ABPs from
different taxa (Hwang et al. 1997; Karn and Nachman
1999; Karn and Laukaitis 2003). We had to take a
different approach here because we wanted to compare potential selection on coding regions (exons) and
noncoding regions (introns), and Ka/Ks have no
meaning for noncoding regions. Instead, we calculated distances, in terms of nucleotide divergences. In
most proteins, the divergence in the coding regions is
expected to be less than that in the noncoding regions
because of purifying (negative) selection acting on the
amino acid sequence of the gene product (thus, there
is less constraint expected on noncoding regions, i.e.,
introns [Hughes and Yeager 1997]). Positive selection
for changes in protein sequence, on the other hand,
could create a situation in which the divergence in
the coding region is actually greater than in the
noncoding regions.
We observed no divergence at all in the coding
regions for the signal peptides. The divergences for
the coding regions of Exons 2 and 3, on the other
hand, are significantly higher (P < 0.01; one-tailed
t-test with infinite degrees of freedom) than those for
either intron. This suggests that positive selection
has acted strongly on the coding regions for the secreted proteins, whereas perhaps the coding regions
for the signal peptides have been under purifying selection. An alternative explanation is that gene conversion was responsible for the conservation of Exon
1 (5¢ UTR and signal peptide). We feel that this is a
less likely explanation because it would have had to
produce the same signal peptide for both the Abpb
and Abpg in both the domesticus and musculus
subspecies of M. musculus (Karn and Laukaitis 2003).
Figure 6 also shows a comparison of the nucleotide divergences for the 5¢ and 3¢ UTRs in Exons 1
and 3, respectively, and for the four helical regions
which are found primarily in Exon 2. Exon 3 contains the rearward portion of Helix 4 and a long tail
not found on most other secretoglobins (Karn and
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
Discussion
We have described three distinct genes, Abpa, Abpb,
and Abpg, encoding the three mouse salivary ABP
subunits, alpha, beta, and gamma, respectively, and
we have produced a physical map for them on the
proximal end of Chr 7. The orientation and proximity of Abpb and Abpg, as well as their nearly
identical intron and exon structures, reinforce the
notion that they are the products of relatively recent
gene duplication (Karn and Laukaitis 2003). Abpa is
probably a more distant relative of these two genes,
in light of its reverse orientation and the different
intron and exon structures. The existence of numerous repeats surrounding Abpa and Abpb and
between Abpb and Abpg suggests that the cluster of
ABP genes could have resulted from one or more
gene duplication events. There is preliminary evidence that additional paralogues of Abpa exist
within the mouse genome (Waterston et al. 2002),
and we have noted additional Abpb/Abpg paralogues. The presence of flanking repeats with
homology to LINE elements suggests that retrotransposons have been actively shaping this area of
Chr 7; work addressing this issue is ongoing.
Abpa and Abpb are orientated with their 5¢ ends
nearest each other and their 3¢ ends running in opposite directions. Thus, these genes share a common
8 kb of upstream DNA. This suggests that they
could be coordinately regulated by a common upstream element. While Abpb and Abpg are 66 kb
apart, they are oriented in the same direction. It
seems possible, therefore, that the expression of
Abpg and Abpb could also have elements in common, e.g., both activated by an enhancer between
Abpa and Abpb.
The secretoglobin family of proteins includes
more than a dozen members found in numerous species of mammals (for a review, see Mukherjee and
Chilton 2000). No function is certain for any of the
secretoglobin family members, but the frequency
with which these secretory proteins are expressed in
tissues which connect the animal’s external environment to its internal physiology suggests that they may
play a role in protecting surfaces (Dominguez 1995).
Gene knockout studies of the best-known protein in
this family, uteroglobin, have had mixed results with
C HR 7
689
one strain developing an IgA nephropathy (Zhang et al.
2000) and the other strain exhibiting increased susceptibility to oxidizing damage to the lung (Stripp et al.
2000). Both of these possible functions are supported
by studies of naturally occurring human uteroglobin
polymorphisms (Choi et al. 2000; Menegatti et al.
2002; Narita et al. 2002; Niimi et al. 2002).
Karn and Laukaitis (2003) constructed a phylogenetic tree with five groups of secretoglobins similar to those described by Ni et al. (2000) but adding
the mouse salivary ABP beta and gamma sequences
and mouse lachrymal and 11 kDa proteins (see Table
1 for the members of groups A–E). Many of the human secretoglobin genes, those representing groups
A, C, and D in the phylogenetic tree, have been
mapped to a region on Chr 11q12.2 (Ni et al. 2000).
Previous work, however, had mapped the class C and
D proteins to Chrs 10q23 and 15q12-q13 (Lehrer et al.
2000) and a uteroglobin-related protein (UGRP1) to
5q31-32 (Niimi et al. 2001).
Murine secretoglobin homologues do not appear
to be concentrated into a single gene cluster, and we
propose the existence of at least three clusters of secretoglobin homologues within the mouse genome.
The uteroglobin cluster of human genes was previously identified on human Chr 11 (Ni et al. 2000) in a
region corresponding to mouse Chr 19 where the
mouse uteroglobin and a potential mouse lipophilin
and mammoglobin have been assigned. A homologue
of the human UGRP1 gene on Chr 5 is found on
mouse Chr 18. The region of mouse Chr 7 where
Abpa and Abpg have mapped genetically (Dlouhy et
al. 1987) and where we have now physically located
them is syntenic to the region of human Chr 19
where the glucose phosphate isomerase (GPI) gene
maps. Also within this region, approximately 1 Mb
closer to the centromere, is a gene homologous to the
major allergen-like 1 (Mjal-1) gene. No human salivary protein equivalent to mouse ABP has yet been
described (Dlouhy et al. 1986).
Interestingly, the secretoglobins described as
classes B (Abpa and Mjal-1) and E (Abpb, Abpg, and
the lachrymal 11-kDa protein; Karn and Laukaitis
2003), which have no identified human homologues,
are located together in the mouse Chr 7 cluster. This
is not entirely surprising since other regions of the
murine genome have duplicated since divergence of
mouse and human from a common ancestor 65–75
million years ago (Waterston et al. 2002). The intriguing question this raises is whether the ABP gene
cluster was duplicated in the mouse and remained
because it serves a unique function, or whether it
was present in the common ancestor and eliminated
from the human line because it served no function
there (Smith et al. 1999).
Our comparison of secretoglobin gene structures
(Table 1) reveals interesting relationships in the sizes
of the three exons and two introns of the 18 secretoglobins for which sequence data are available.
Secretoglobin groups A–E were derived by molecular
evolutionary comparisons of amino acid sequences,
and yet the sizes of the exons and introns of their
genes that we present in Table 1 reinforce that evolutionary grouping. Evolutionary considerations lead
one to predict that the exon size would be more
strongly conserved than intron size, since the exons
contain the coding regions, and this prediction is
realized. Variation in gene size among the five
groups of secretoglobins is primarily influenced by
variability in intron size, as shown by the relatively
large standard deviations from the mean intron sizes
in each of the five groups.
Further, one would predict that the first and third
exons would be more variable in size than the middle
exon because they contain the 5¢ and 3¢ UTRs which
are also noncoding sequences, and this prediction too
is realized. In fact, the size of Exon 2 is maintained at
188 bp in all but a few cases, most notably cluster E.
That group consistently lacks the first three nucleotides and the last three nucleotides of Exon 2. The effect on the ABP beta and gamma subunits is to extend
their signal peptides by one amino acid on the C-terminus and to reduce the length of each secreted subunit by one amino acid. Group E also has the cleavage
point of the signal peptide shifted to make the first Cys
in the amino acid sequence residue 1 in the secreted
protein, whereas it is residue 3 in all the other secretoglobins. Whether the change in the size of the
middle exon in the group E genes is related to the shift
in the cleavage point for the signal peptide is unclear.
One of the most interesting features of the ABP
system is the strong Darwinian selection that has been
shown to be operating on one or more of these genes.
Several studies of Abpa have suggested that its evolution has deviated significantly from the predictions
of the Neutral Theory of Molecular Evolution (Kimura
1983), probably as the result of strong directional selection (Hwang et al. 1997; Karn and Nachman 1999;
Karn et al. 2002). It has been proposed that selection on
Abpa has proceeded by selective sweeps in at least two
subspecies of Mus musculus: M. m. domesticus (Karn
and Nachman 1999) and M. m. musculus (Karn et al.
2002). More recently, Karn and Laukaitis (2003) have
suggested that directional selection is probably also
driving the evolution of Abpb and Abpg. This is
probably owing to selection directly on these genes,
rather than hitchhiking, since the strongest evidence
is provided by a relatively high nonsynonymous/synonymous substitution rate ratio (Ka/Ks) as shown for
Abpa (Hwang et al. 1997; Karn and Nachman 1999)
C.M. L AUKAITIS
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
690
C.M. L AUKAITIS
and for Abpb and Abpg (Karn and Laukaitis 2003). In
the case of Abpa, the gene encoding the alpha subunit,
the one common to both dimer forms (A:B and A:G),
there is evidence that the directional selection may
take the form of sexual selection (Talley et al. 2001;
Laukaitis et al. 1997). It remains to be seen whether
sexual selection might also explain the relatively high
Ka/Ks ratios observed for Abpb and Abpg.
The two different ABP dimers, A:B and A:G, have
been shown to bind testosterone and dihydrotestosterone (DHT) with different affinities (Karn and Clements 1999); this has been attributed to differences in
key residues in the hydrophobic pockets of the two
dimers (Karn and Laukaitis 2003). The role of ligand
binding in the function of ABP and, for that matter, the
other secretoglobins, is unknown. It is likely, however, that the evolution of Abpb and Abpg since their
divergence from a common ancestral sequence has
proceeded under the influence of a need to bind different ligands (Karn and Laukaitis 2003). This idea is
supported by the significantly higher nucleotide divergences in the coding region (exons 2 and 3) for the
secreted proteins than in their introns or the coding
region for their signal peptides. We suggest that the
evolutionary pressure that likely fixed the duplication
of the gene ancestral to Abpb and Abpg and resulted in
the expression of the duplicated products is a selection
pressure that is not likely to be influencing the evolution of Abpa because its product is common to both
dimers. An alternative explanation to our view that
higher nucleotide divergences reflect positive selection on exons 2 and 3 of Abpb and Abpg is that mutation rates could vary between exons and introns. It
seems unlikely (though not impossible) that mutation
rates vary in such a nonrandom way, but it is quite
likely that the amount of constraint influencing nucleotide divergence on gene regions is different (Hughes and Yeager 1997). In other words, the high
divergence at exons compared with introns might be
driven by positive selection on exons, unusually
strong negative (purifying) selection on introns, or
some combination of both. Since there is no evidence
that the introns of Abpb and Abpg are under any
constraint different from the introns of other genes
that have been studied thus far, we favor the view that
our data support positive selection on the exons.
Closer analysis, in which the nucleotide divergences of the coding regions of Abpb and Abpg (signal
peptide, the four helices, and the tail) are compared
with each other and with the noncoding regions (5¢
and 3¢ UTRs and the introns), creates an even more
intriguing picture. The lower level of divergence in
the noncoding regions and in two parts of the coding
region, the signal peptide and Helix 3, suggests that
the duplication that produced Abpb and Abpg may
not be very old. This is because the nucleotide divergence of Helix 3 is not significantly different from
the noncoding regions, which are not very high. By
contrast, the values for the other three helices are
significantly higher. It appears that positive selection
drove the divergence of Helices 1, 2, and 4. Helix 3, by
contrast, was not under the same pressure, and the
signal peptide may even have been conserved by purifying selection. Karn and Laukaitis (2003) suggested
that the conservation of Helix 3 between the beta and
gamma subunits may be due to a critical role in the
dimer interface where the ligand is bound (Callebaut
et al. 2000), which may explain why it has been
conserved relative to the other helices which contain
residues that may control the specificity for the ligand bound (Karn and Laukaitis 2003).
The possibility of selection occurring directly on
one or more of the genes in the ABP cluster raises the
interesting question of the effect this may have had
on the evolution of other DNA sequences within and
around the cluster. This begs the production of sequences of a large region of this chromosome segment in other species of Mus as well as in the three
subspecies of Mus musculus. Such a comparison
would shed light on a number of very interesting
issues, such as the effects of selection on the ABP
genes and their surrounding sequences and the
question of how recently duplications producing
ABP gene paralogues occurred.
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
C HR 7
Acknowledgments
The authors thank M. Nachman and R. Emes for
helpful discussions and advice. This work was supported in part by the Holcomb Research Institute,
Butler University, and that support is gratefully acknowledged.
References
3 1. Agarwal S, Viola JP, Rao A (1999) Chromatin-based
regulatory mechanisms governing cytokine gene transcription. J Allergy Clin Immunol 103, 990–999
2. Altschul SF, Gish W, Miller W, Myers DE, Lipman DJ
(1990) Basic local alignment search tool. J Mol Biol 215,
403–410
3. Callebaut I, Poupon A, Bally R, Demaret JP, Housset D
et al. (2000) The uteroglobin fold. Ann N Y Acad Sci
923, 90–112
4. Choi M, Zhang Z, Ten Kate LP, Collee JM, Gerritsen J
et al. (2000) Human uteroglobin gene polymorphisms
and genetic susceptibility to asthma. Ann N Y Acad
Sci 923, 303–306
5. Dlouhy S, Karn R (1983) The tissue source and cellular
control of the apparent size of androgen binding protein
(Abp), a mouse salivary protein whose electrophoretic
C HR 7
691
mobility is under the control of Sex-limited saliva
pattern (Ssp). Biochem Genet 21, 1057–1070
Dlouhy S, Karn R (1984) Multiple gene action determining a mouse salivary protein phenotype: identification of the structural gene for Androgen binding
protein (Abp). Biochem Genet 22, 657–667
Dlouhy R, Nichols W, Karn R (1986) Production of an
antibody to mouse salivary androgen binding protein
(ABP) and its use in identifying a prostate protein produced by a gene distinct from Abp. Biochem Genet 24,
673–743
Dlouhy S, Taylor B, Karn R (1987) The genes for mouse
salivary Androgen-binding protein (ABP) subunits alpha and gamma are located on chromosome 7. Genetics 115, 535–543
Dominguez P (1995) Cloning of a Syrian hamster
cDNA related to sexual dimorphism: establishment of
a new family of proteins. FEBS Lett 376, 257–261
Graur D, Li W-H (2000) Fundamentals of Molecular
Evolution. (Sunderland, Mass.: Sinauer Associates, Inc.)
Hubbard T, Barker D, Birney E, Cameron G, Chen Y et
al. (2002) The Ensembl genome database project. Nucleic Acids Res 30, 38–41
Hughes AL, Yeager M (1997) Comparative evolutionary rates of introns and exons in murine rodents. J Mol
Evol 45, 125–130
Hwang J, Hoffstetter J, Bonhomme F, Karn R (1997) The
microevolution of mouse salivary androgen-binding
protein (ABP) paralleled subspeciation of Mus musculus.
J Hered 88, 93–97
Karn R, Clements M (1999) A comparison of the structures of the alpha:beta and alpha:gamma dimers of mouse
salivary androgen-binding protein (ABP) and their differential steroid binding. Biochem Genet 37, 187–199
Karn R, Laukaitis C (2003) Characterization of two
forms of mouse salivary androgen-binding protein
(ABP): Implications for evolutionary relationships and
ligand-binding function. Biochemistry 42, 7162–7170
Karn R, Nachman M (1999) Reduced nucleotide variability at an androgen-binding protein locus (Abpa) in
house mice: evidence for positive natural selection.
Mol Biol Evol 16, 1192–1197
Karn R, Russell R (1993) The amino acid sequence of
the alpha subunit of mouse salivary androgen-binding
protein (ABP), with a comparison to the partial sequence of the beta subunit and to other ligand-binding
proteins. Biochem Genet 31, 307–319
Karn R, Orth A, Bonhomme F, Boarsot P (2002) The
complex history of a gene proposed to participate in a
sexual isolation mechanism in house mice. Mol Biol
Evol 19, 462–471
Karolchik D, Baertsch R, Diekhans M, Furey TS, Hinrichs A et al. (2003) The UCSC genome browser database. Nucleic Acids Res 31, 51–54
Kent WJ (2002) BLAT–the BLAST-like alignment tool.
Genome Res 12, 656–664
Kimura M (1983) The Neutral Theory of Molecular Evolution. (Cambridge, U.K.: Cambridge University Press)
Klug J, Beier HM, Bernard A, Chilton BS, Fleming TP
et al. (2000) Uteroglobin/Clara cell 10-kDa family of
proteins: nomenclature committee report. Ann N Y
Acad Sci 923, 348–354
Klug WS, Cummings MR (2003) Concepts of Genetics.
Upper Saddle River, NJ; Prentice Hall
Kratz E, Dugas JC, Ngai J (2002) Odorant receptor gene
regulation: implications from genomic organization.
Trends Genet 18, 29–34
Kumar S, Tamura K, Jakobsen I, Nei M (2001) Mega2:
Molecular Evolutionary Genetics Analysis Software.
Bioinformatics 17, 1244–1245
Laukaitis C, Critser E, Karn R (1997) Salivary androgen-binding protein (ABP) mediates sexual isolation in
Mus musculus. Evolution 51, 2000–2005
Lehrer RI, Nguyen T, Zhao C, Ha CX, Glasgow B (2000)
Secretory lipophilins: a tale of two species. Ann N Y
Acad Sci 923, 59–67
Menegatti E, Nardacchione A, Alpa M, Agnes C, Rossi
D et al. (2002) Polymorphism of the uteroglobin gene
in systemic lupus erythematosus and IgA nephropathy.
Lab Invest 82, 543–546
Mukherjee A, Chilton B eds. (2000) The uteroglobin/
Clara cell protein family. Ann N Y Acad Sci 923
Narita I, Saito N, Goto S, Jin S, Omori K et al. (2002)
Role of uteroglobin G38A polymorphism in the progression of IgA nephropathy in Japanese patients. Kidney Int 61, 1853–1858
Ni J, Kalff-Suske M, Gentz R, Schageman J, Beato M
et al. (2000) All human genes of the uteroglobin family
are localized on chromosome 11q12.2 and form a dense
cluster. Ann N Y Acad Sci 923, 25–42
Niimi T, Keck-Waggoner CL, Popescu N, Zhou Y, Levitt
B et al. (2001) UGRP1, a uteroglobin/Clara cell secretory
protein-related protein, is a novel lung-enriched downstream target gene for the T/EBP/NKX2.1 homeodomain
transcription factor. Mol Endocrinol 15, 2021–2036
Niimi T, Munakata M, Keck-Waggoner C, Popescu N,
Levitt B et al. (2002) A polymorphism in the human
UGRP1 gene promoter that regulates transcription is
associated with an increased risk of asthma. Am J Hum
Genet 70, 718–725
Smith NG, Knight R, Hurst L (1999) Vertebrate genome evolution: a slow shuffle or a big bang? Bioessays
21, 697–703
Stripp B, Reynolds S, Plopper C, Boe I, Lund J et al.
(2000) Pulmonary phenotype of CCSP/UG deficient
mice: a consequence of CCSP deficiency or altered
Clara cell function? Ann N Y Acad Sci 923, 202–209
Talley H, Laukaitis C, Karn R (2001) Female preference for male saliva: implications for sexual isolation of Mus musculus subspecies. Evolution 55,
631–634
Waterston R, Lindblad-Toh K, Birney E, Rogers J, Abril J
et al. (2002) Initial sequencing and comparative analysis of the mouse genome. Nature 420, 520–562
White R (2001) Gene Transcription: Mechanisms and
Control. (London, U.K.: Blackwell Science Ltd.)
Zhang Z, Kundu G, Zheng F, Yuan C, Lee E et al. (2000)
Insight into the physiological function(s) of uteroglobin
by gene-knockout and antisense-transgenic approaches. Ann N Y Acad Sci 923, 210–233
C.M. L AUKAITIS
4
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
5
21.
22.
6
ET AL .:
M OUSE ABP G ENE C LUSTER
ON
23.
24.
25.
26.
27.
28.
29.
7
30.
31.
32.
33.
8
34.
35.
36.
37.
38.
39.