Genomics 56, 303–309 (1999)
Article ID geno.1998.5721, available online at http://www.idealibrary.com on

The Genomic Organization of Type I Keratin Genes in Mice
Hajime Sato,* ,† Tsuyoshi Koide,† Tomoko Sagai,† Sei-Ichi Ishiguro,* Makoto Tamai,*Naruya Saitou,‡ and Toshihiko Shiroishi† ,1*Department of Ophthalmology, Tohoku University School of Medicine, Seiryo-Machi 1-1, Sendai, Miyagi-ken 980-8574, Japan;†Mammalian Genetics Laboratory and ‡Laboratory of Evolutionary Genetics, National Institute of Genetics,
Yata-1111, Mishima, Shizuoka-ken 411-8540, Japan
Received July 31, 1998; accepted December 7, 1998

We isolated two new keratin cDNAs by screening a
cDNA library constructed from poly(A) 1 RNA of thedorsal and abdominal skin of C57BL/10J mice with aprobe of human KRT14. Due to its high sequence homology to human keratin 17 cDNA, one full-length
cDNA is most likely to be mouse keratin 17 (Krt1-17)
cDNA. The other is the putative full-length cDNA of anovel type I keratin gene, designated Krt1-c29. These
two keratin genes were mapped to the distal portion of
Chromosome 11, where the mouse keratin gene complex-1 (Krt1) is localized. To elucidate the genomic
organization of Krt1 in mice, we carried out genetic
and physical analyses of Krt1. A large-scale linkage
analysis using intersubspecific backcrosses suggested
that there are two major clusters in Krt1, one containing Krt1-c29, Krt1-10, and Krt1-12 and the other containing Krt1-14, -15, -17, and -19. Truncation experiments with two yeast artificial chromosome clones
containing the two clusters above have revealed thatthe gene order of Krt1 is centromere–Krt1-c29 –Krt110 –Krt1-12–Krt1-13–Krt1-15–Krt1-19 –Krt1-14 –Krt1-17–
telomere. Finally, we analyzed sequence divergence
between the genes belonging to the Krt1 complex. The
results clearly indicated that genes are classified into
two major groups with respect to phylogenetic relationship. Each group consists of the respective genecluster demonstrated by genetic and physical analyses
in this study, suggesting that the physical organization of the Krt1 complex reflects the evolutionary process of gene duplication of this complex. © 1999 Academic
Press

INTRODUCTION

Keratins are a group of water-insoluble proteins constituting paired acidic and basic proteins that form
intermediate filaments (IF) in epithelial cells. Coordinated expression of the acidic and basic proteins, which
are encoded by the type I and type II keratin genecomplexes, is essential for normal epithelial integrity.
1
To whom correspondence should be addressed. Telephone: 81559-81-6818. Fax: 81-559-81-6817. E-mail: tshirois@lab.nig.ac.jp.

Characterization of the genomic organization is crucial
for a better understanding of the regulation of keratin
genes and the etiology of many genetic defects known
to cause abnormal development of epithelial cells.
According to immunoreactivity and sequence homologies as well as relative charge, K10, K11, K12, K13,
K14, K15, K16, K17, K18, and K19 belong to type I
keratin, while K1, K2, K3, K4, K5, K6, K7, and K8
belong to type II keratin (Moll et al., 1982; Eichner etal., 1984). As is the case with other IF, a keratin chain
consists of an amino-terminal end domain, an a-helicalrod domain, and a carboxyl-terminal end domain. In
each group, the central a-helical rod domain is remarkably conserved in its size, secondary structure, andsequence, while the amino- and carboxyl-terminal end
domains differ in size and sequence (Steinert and Roop,
1988). Specific pairs of at least one type I and one type
II keratin protein are coexpressed in a tissue- anddifferentiation-specific manner and are required for IF
assembly. K1, K2, K5, K10, and K14 are expressed innormal epidermis; K3 and K12 are expressed in cornea;
K4, K13, and K15 are expressed in esophagus andother nonkeratinizing epithelia; K8 and K18 are expressed in simple epithelia; K19 is expressed in periderm; and K9 is expressed in palmar and plantar tissue. K6, K16, and K17 are expressed mainly in theepidermis under pathological hyperproliferative states(Blumenberg, 1993).
It is reported that mutations in keratin genes cause
many hereditary human diseases (Coulombe et al.,
1991; Cheng et al., 1992; Reis et al., 1994; McLean etal., 1995; Richard et al., 1995; Rugg et al., 1995; Irvine
et al., 1997). Mutant keratin generates the abnormal
keratin filament network that is visible as perinuclear
keratin aggregates with keratin clumps. The disruption of the keratin network causes keratinocytes to be
fragile, as was demonstrated in transgenic mice andknockout mice (Vassar et al., 1991; Kao et al., 1996).
Further study of the keratin genes will be useful for
diagnosis of inherited diseases of epithelia.
In humans, the type I keratin complex has been
mapped to chromosome 17q12– q21. Recently, it was

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and ligated into the EcoRI site of Lambda ZAP II vector (Stratagene,
La Jolla, CA). The ligated mixture was packaged with Gigapack II
Gold Packaging Extract (Stratagene).
Screening procedures and cDNA cloning. The cDNA library was
screened by plaque hybridization with 32P-labeled human keratin 14
as a probe by standard methods (Sambook et al., 1989). The DNA
fragment of KRT14 was amplified by polymerase chain reaction(PCR) using specific primers (KRT14-F, 5-ATGACTACCTGCAGCCGCCA-3; KRT14-R, 5-TTGATCTCAGCAGGCCGCTG-3) (Marchuk
et al., 1985). Washing was performed in 23 SSC, 0.5% sodium
dodecyl sulfate (SDS) for 1 h at room temperature followed by 0.13
SSC, 0.25% SDS for 10 –15 min at 65°C. Seventeen independent
clones were further analyzed.
DNA sequence analysis. The DNA sequence was determined forboth strands by a 377 automated DNA sequencer (Perkin–Elmer
Applied Biosystems, CT) using a dRhodamine Terminator Cycle
Sequencing FS Ready Reaction Kit (Perkin–Elmer Applied Biosystems) and analyzed by DNASIS version 3.0 (Hitachi Software, Japan). Homologous sequences were searched for in the database at the
National Center for Biotechnology Information (NCBI) using the
BLAST network service.

FIG. 1. Comparison of the amino acid sequences of mouse
Krt1-17 and human keratin 17 genes. Vertical lines denote identicalamino acid residues. Note that the percentage of identical amino acidresidues is 95%. The rod domain of the keratin chain consists of threea-helical tracts termed Coil 1A, Coil 1B, and Coil 2, interrupted bylinker 1 and linker 12. The boundary of these domains is based on the
database. MK17, mouse keratin 17; HK17, human keratin 17.

reported that the genes described below are organizedwithin a 55-kb DNA fragment in the following
order: 59– KRT19 –KRT15–KRT17–KRT16 –KRT14–39(Milisavljevic et al., 1996). In mice, the type I keratin
complex (Krt1) was mapped to the distal portion of
Chromosome 11, which is syntenic to human chromosome 17q12– q21. Although Krt1-19, Krt1-15, andKrt1-13 are separated by 5– 6 kb each and are in thesame transcriptional orientation (Nozaki et al., 1994;
Filion et al., 1994), the complete genomic organizationof Krt1 has not yet been determined.
In this study, we have isolated mouse Krt1-17 and anovel type I keratin cDNA (Krt1-c29) from a skin cDNA
library. We determined the comprehensive gene orderof Krt1 by fine linkage analysis and the physical organization of Krt1 by analysis of YAC clones. Finally, we
discuss the phylogenetic relationship of the cloned Krt1
genes.

Northern blot analysis. Seven micrograms each of total RNA
derived from several tissues was electrophoresed on a 1.2% formaldehyde gel by a standard method (Sambrook et al., 1989) and blotted
onto Hybond-N 1 nylon membrane (Amersham Pharmacia Biotech,
Buckinghamshire, UK). Hybridization was carried out with a 32Plabeled 628-bp EcoRI–PvuII cDNA fragment of clone pKS29. Washing was performed at room temperature in 23 SSC, 0.5% SDS for 1 h
and subsequently at 60°C in 0.13 SSC, 0.25% SDS for 10 min.
Autoradiographs were analyzed with the BAS 2000 Bioimage Analyzer (Fuji Photo Film, Tokyo, Japan).
Linkage analysis. Linkage analyses were carried out using threedifferent sets of mouse DNA panels previously constructed (Sagai etal., 1998; Sato et al., 1998). Microsatellite markers were purchasedfrom Research Genetics (Huntsville, AL). The sequences of oligonucleotide primers for Krt1-10 and Krt1-12 were described previously(Sato et al., 1998). The oligonucleotide primers set for Krt1-14, Krt115, Krt1-17, Krt1-19, and Krt1-c29 were as follows: Krt1-14F, 59AGATCCGCACCAAGGTCATGG-39 and Krt1-14R, 59-GTGCAACTCAGAAAAAGAAGC-39. The Krt1-14 39-noncoding region probe wasobtained from Dr. Dennis Roop (Knapp et al., 1987). The sequencesfor Krt1-15F, 59-GAGAGAAATCTAAGTGTCTGGTG-39, and Krt115R, 59-GATAACACCCTTTTAGAACTGG-39 (Nozaki et al., 1994),
Krt1-17F, 59-ACCACCCGTTAAGGACTCAG-39, and Krt1-17R, 59CCATGGTCATTTATTTCAGTG-39, were obtained from analyses of
pKS11, pKS14, pKS17, pKS18, pKS20, and pKS28 clones in thisstudy and the published sequence (Knapp et al., 1987). Krt1-19F,
59-CCAAGGCCATCTGAGCTACC-39, and Krt1-19R, 59-CCTTGGAGGATAGTTTTATTGAC-39 (Lussier et al., 1990), Krt1-c29F, 59ACAACGCCAACATCATCCTG-39, and Krt1-c29R, 59-TCTTGAGGTATGCCAGCTCC-39, were obtained from the analysis of the pKS29
clone in this study. Simple sequence-length polymorphism and sin-

MATERIALS AND METHODS
Construction of the skin cDNA library. Total RNA of dorsal andabdominal skin of adult C57BL/10J mice purchased from The Jackson Laboratory (Bar Harbor, ME) was extracted with guanidinium
thiocyanate by a standard protocol (Sambrook et al., 1989). Polyadenylated RNA was purified from the total RNA with a mRNA Purification Kit (Amersham Pharmacia Biotech, Uppsala, Sweden).
cDNA was synthesized from 5 mg of mRNA with a cDNA synthesis
module (Amersham Pharmacia Biotech, Buckinghamshire, UK) using anchored dT25 primers. The cDNA was methylated with EcoRI
methylase and ligated with 59-phosphorylated EcoRI linker. After
digestion with EcoRI, it was size-fractionated with SizeSepTM400
Spun Columns (Amersham Pharmacia Biotech, Uppsala, Sweden)

FIG. 2. Northern blot of Krt1-c29. The Krt1-c29 probe detected
two signals of approximately 1.5 and 3.5 kb in skin. The intensity ofsignals suggested that the 1.5-kb mRNA is the major transcript.
G3PDH is the internal control. Lanes 1, whole embryo (11.5 days
postcoitus (dpc); 2, whole embryo (12.5 dpc); 3, placenta (12.5 dpc); 4,
liver (adult); 5, testis (adult); 6, skin (adult); 7, lung (adult).

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305

FIG. 3. Comparison of the amino acid sequences of type I keratin rod domains. The amino acid sequences are optimally aligned. Dashes
denote deletions introduced to obtain optimal alignment. White letters with a black background represent identical amino acid residues.
There is 70 – 80% identity among the genes. XEN.47KD, Xenopus laevis 47-kDa type I keratin; BOV.VIB, bovine type I keratin VIB;
MK59KD, mouse 59-kDa type I keratin; MK, mouse keratin.
gle-strand conformation polymorphism for Krt1-10, Krt1-15, Krt1-19,
and Krt1-c29 were performed as previously described (Sato et al.,1998). The PCR products amplified with Krt1-14 primers were visualized by staining with ethidium bromide by 16% polyacrylamide gelelectrophoresis. The linkage analysis was performed with Map Manager v2.6.5 and the QTB11 program (Manly, 1993).

of YACs, 1993). Agarose plugs were loaded onto a 1% gel and sizefractionated in a CHEF Mapper XA Pulsed Field Electrophoresis
System (Bio-Rad). Gels were blotted onto Hybond-N 1 nylon membrane, and Southern hybridization was performed with a 32P-labeled
pBR322 probe that hybridizes with the left arm of the YAC vector.
Washing and autoradiographs were performed as described above.

Isolation of the yeast artificial chromosome (YAC) clone. A
genomic YAC library (Research Genetics) was screened by PCR withspecific primers for each keratin gene. Clone YAC120F10 (approximately 430 kb), clone YAC185F10 (approximately 530 kb), and clone
YAC112A6 (approximately 580 kb) were isolated and further analyzed.

Phylogenetic tree analysis. The neighbor-joining method (Saitou
and Nei, 1987) was used for constructing phylogenetic trees of mouse
type I keratin genes. CLUSTAL W (Thompson et al., 1994) was used
for multiple alignment and tree construction.

Truncation of YAC clones. Truncation vectors pB1F and pB1R,
which carry a mouse B1 repeat sequence in different orientations, acopy of the yeast lysine-2 (Lys2) gene, and the telomere sequence,
were kindly provided by J. Edmondson and R. Rothstein. Isolated
YAC clones were transformed with pB1F or pB1R vector, which was
linearized by digestion with SalI using an alkali cation yeast transformation kit (Bio 101, Inc., CA), and transformed yeast cells wereplated on synthetic complete (SC) medium lacking tryptophan andlysine. After 2 days, colonies were plated in duplicate on SC (Lys-,
Trp-) and SC (Lys-, Ura-) plates. Those colonies that grew only on SC(Lys-, Trp-) plates were further analyzed (Lewis et al., 1992).
Pulsed-field gel electrophoresis and Southern blot analysis. Selected colonies were examined by PCR with the above primers todetermine whether there were any Krt1 genes and microsatellite
markers. The sequences of oligonucleotide primers for Krt1-13 wereas follows: Krt1-13F, 59-GTTTCTCCAGCCTAAGCACC-39; Krt113R, 59-TCCAATCAGAAGAGATGGGG-39 (Filion et al., 1994). DNA
from successfully truncated YAC clones was prepared in low-melting-point agarose (SeaPlaque GTG agarose; FMC BioProducts, Rockland, ME) according to the protocol provided at the training course atCold Spring Harbor Laboratory (Analysis and Genetic Manipulation

RESULTS

Isolation of Two New Type I Keratin cDNAs
In this study, we isolated two type I keratin cDNAs
from a skin cDNA library. One full-length cDNA washighly homologous to human keratin 17 cDNA (Troyanovsky et al., 1992) and was designated mouse keratin 17 (Krt1-17) cDNA. The complete nucleotide sequence of the Krt1-17 cDNA was determined by the
analysis of six independent clones, pKS11, pKS14,
pKS17, pKS18, pKS20, and pKS28. This sequence hasan open reading frame (ORF) of 1299 bp and encodes aprotein of 433 amino acids (DDBJ Accession No.
AB013608). The deduced amino acid sequence is 95.4%
identical with human keratin 17 over 433 amino acids(Fig. 1). The second was the cDNA of a novel keratin
gene, which we designated Krt1-c29. Its nucleotide sequence was determined by analysis of the clone pKS29.

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FIG. 4. Linkage analysis of mouse type I keratin genes. Recombination distances between loci in centimorgans are shown to the left ofthe chromosome, and microsatellite markers and gene loci are shown to the right. Note that there are two clusters in Krt1, one containing
Krt1-c29, Krt1-10, and Krt1-12 and the other containing Krt1-14, -15, -17, and -19. Rim3, recombination-induced mutation 3; Rara, retinoic
acid receptor a; Grn, granulin; Jup, junctional plakoglobin.

It also has a long ORF of 1344 bp and encodes a proteinof 448 amino acids (DDBJ Accession No. AB013607).
Northern blot analysis demonstrated the presence of amajor 1.5-kb transcript in adult mouse skin but not inany other tissues examined (Fig. 2). Additionally, thededuced amino acid sequence in the keratin rod domain has 70 – 80% identity with other type I keratin
genes (Fig. 3).
Linkage Analysis of Mouse Type I Keratin Genes
We carried out a linkage analysis of Krt1-17 andKrt1-c29 and other mouse type I keratin genes, basedon three different crosses. Krt1-17 and Krt1-c29 were
mapped to the distal portion of mouse Chromosome 11,
where other Krt1 genes are located (Fig. 4). The linkage analysis in Cross I revealed that Krt1-c29 is located proximal to other Krt1 genes. In Cross II, Krt1-10
and Krt1-12 were segregated from Krt1-15, Krt1-19,
Krt1-14, and Krt1-17 and mapped proximal to them.
The result of Cross III showed that Krt1-15 is locatedproximal to the Krt1-19, Krt1-14, and Krt1-17 genes.
Finally, these linkage analyses indicated that the geneorder of Krt1 is centromere–Krt1-c29–(0.81 cM)– Krt110, Krt1-12–(0.42 cM)–Krt1-15–(0.13 cM)–Krt1-19,
Krt1-14, Krt1-17–telomere.

vector. Physical separation of Krt1-c29, Krt1-10, andKrt1-12 by the truncation of YAC120F10 (Fig. 5) indicated that they are localized in following order: centromere–Krt1-c29 –Krt1-10 –Krt1-12–telomere and suggested that the distance between Krt1-c29 and Krt1-10
is within 80 kb and that Krt1-10 is located within 40 kb
of Krt1-12. Additionally, separation of Krt1-19, Krt114, and Krt1-17 by the truncation of YAC185F10 (Fig.
5) indicated that they are localized in the following
order: centromere–Krt1-13, Krt1-15–Krt1-19 –Krt1-14 –Krt1-17–telomere and suggested that Krt1-19 andKrt1-14 are separated by a distance of 20 –100 kb andKrt1-17 is situated within a distance of 130 kb from
Krt1-14. The failure to separate Krt1-13, Krt1-15, andKrt1-19 suggested that these genes are tightly clustered, which is consistent with previous reports (Filion
et al., 1994). The result of the truncation of YAC112A6
was the same as that of YAC185F10 except for the
distance between Krt1-19 and Krt1-14. The distance
between Krt1-19 and Krt1-14 was estimated to be 20 –100 kb in YAC185F10 and 100 –180 kb in YAC112A6.
This difference is possibly due to an artifactual deletion in the clone YAC112A6. The physical organizationof the Krt 1 complex is summarized in Fig. 6.

Physical Organization of Mouse Type 1 Keratin Genes

Phylogenetic Tree Analysis of Mouse Type I Keratin
Genes

To determine the gene order in Krt1 independently,
YAC truncation was performed using a pB1F or pB1R

We constructed a phylogenetic tree of the genes to
explore the evolution of mouse type I keratin genes

ORGANIZATION OF MOUSE TYPE I KERATIN GENES

307

FIG. 5. Southern hybridization of serial truncated YAC clones. Southern hybridization with a YAC left-arm probe revealed that YAC
120F10 and YAC 185F10 were successfully truncated with pB1F or pB1R. F or R indicates a clone truncated with pB1F or pB1R, respectively.
The lower panels represent the result of PCR. Krt1 genes and microsatellite markers are shown to the left. Lane M, 50-kb lambda laddermarker; Ori, original YAC clone.

(Fig. 7). The result clearly showed that Krt1 genes areclassified into two major groups, one group including
Krt1-10, Krt1-12, and Krt1-c29, and the other including
Krt1-13, -15, -19, -14, and -17. Each member of the twogroups might have arisen from ancestral genes by gene
duplication. The grouping based on this phylogenetic
tree showed a good correlation with the two clusters of
Krt1 genes in the mouse genome.

DISCUSSION

We have presented here the sequence data of two
cDNAs of type I keratin genes, Krt1-17 and Krt1-c29,
which were cloned from a skin cDNA library. It ishighly likely that Krt1-17 is the mouse orthologue ofhuman KRT17, because of the high percentage ofamino acid sequence identity and the similar expres-

FIG. 6. Organization of the mouse Krt1 genes. The gene order and the distance between adjacent genes denoted above and below thechromosome, respectively, are based on the result of the serial truncation of the YAC clones. Those of Krt1-13, -15, and -19 are based on thephysical map of cosmid clones. DNA clones used in this study are shown above the chromosome. Arrows at Krt1-13, -15, and -19 indicate thetranscriptional orientation. R, right arm of YAC; L, left arm of YAC.

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FIG. 7. A phylogenetic tree of eight mouse keratin genes. The
location of the root (designated as a broken line) was estimated byusing the mouse desmin sequence as the outgroup. Numbers on
interior branches are bootstrap probabilities (%). Scale is number ofamino acid substitutions per site.

sion pattern (Knapp et al., 1987). Knapp and co-workers presented data for the sequence of pkSCC50, which
seemed to be a partial cDNA fragment of the mouseorthologue of human KRT17. The sequence of pkSCC50 appears to be identical to a 0.8-kb 39-region of
Krt1-17 cDNA. They also reported that the expression
of kSCC50 was detected in normal hair follicles, hyperproliferative epidermis, and squamous cell carcinoma,
where human KRT17 is expressed as well (Moll et al.,
1982; Troyanovsky et al., 1992). While this article wasin review, another description of mouse Krt1-17 reporting a nucleotide sequence identical to that of ours was
published (McGowan and Coulombe, 1998).
On the other hand, Krt1-c29 is a novel gene thatbelongs to Krt1 as suggested by the alignment of thetype I keratin rod domain (Fig. 3). Northern blot analysis in this study (Fig. 2) demonstrated that the size of
the Krt1-c29 major transcript was approximately 1.5
kb. In addition, since the sequence ACCATGT flanking
the AUG initiator codon of Krt1-c29 cDNA has homology to the Kozak consensus sequence, ACCATGG(Kozak, 1986, 1987), Krt1-c29 is probably a full-length
cDNA. The presence of a minor transcript approximately 3.5 kb in size indicates that there is an alternative promoter or alternative splicing in the Krt1-c29
gene, because genomic Southern hybridization with a
0.6-kb 59-fragment of the Krt1-c29 cDNA as a probegave a unique strong signal.
The high-resolution linkage analyses in this study(Fig. 4) revealed that the gene order in Krt1 is centromere–Krt1-c29 –Krt1-10, Krt1-12–Krt1-15–Krt1-19,
Krt1-14, Krt1-17–telomere and suggested that there
are two major clusters in the Krt1 complex, one containing Krt1-c29, Krt1-10, and Krt1-12 and the othercontaining Krt1-15, Krt1-19, Krt1-14, and Krt1-17.
The mouse type I hair keratin 2 and 3 genes Krt1-4 andKrt1-2 (Winter et al., 1994) did not recombine with
D11Mit123. The oligonucleotide primers forD11Mit123
are set within Krt1-1 (data not shown). In conjunction
with these data, the fact that Krt1-1, Krt1-4, andKrt1-2 are present in YAC185F10 and YAC112A6, but
not in cosmids 32 and 43 (Fig. 6), suggests that themouse type I hair keratin genes are located between

Krt1-12 and Krt1-13. It has been proposed that Krt1
genes are responsible for mouse mutations with skinand hair anomalies, such as Re, Bsk, and Rim3 (Eicher
and Varnum, 1986; Lyon and Zenthon, 1986, 1987;
Nadeau et al., 1989; Liu et al., 1994; Sato et al., 1998).
Linkage analysis in this study, however, indicated that
the Krt1 complex including type I hair keratin genes
are excluded as candidates for Rim3, which may beallelic to Re den.
We have determined the genomic organization of the
Krt1 complex in mice using the YAC truncation method(Fig. 6). The isolation of a clone truncated between
D11Mit59 and Krt1-19 in YAC185F10 (Fig. 5) determined the transcriptional orientation of Krt1-19, because D11Mit59 is located within the first intron of
Krt1-19 and the PCR primer set for Krt1-19 used inthis study is located at the 39 noncoding region. Because there are several B1 repetitive sequences in thefirst intron of Krt1-19, a recombination could occur in
one of them (Lussier et al., 1990). We constructed a
restriction map of two overlapping cosmid clones isolated from a mouse cosmid library (Clontech, Palo Alto,CA). The combined size of the two clones was approximately 55 kb, and they contained Krt1-13, Krt1-15,
and Krt1-19 (data not shown). This is consistent withthe study of Filion et al. (1994) and suggested that
Krt1-14 and Krt1-17 are located at least 5 kb from the
59-region of Krt1-19. Although we need to obtain amore precise physical map and to determine the transcriptional orientation of each gene, it is likely that the
genome organization of Krt1-13, Krt1-15, Krt1-19,
Krt1-14, and Krt1-17 is different from that in the human genome (Milisavljevic et al., 1996). Conversely, it
would be intriguing to analyze the genomic organization of human orthologues of Krt1-c29, Krt1-10, andKrt1-12.
A phylogenetic analysis of the Krt1 complex clearly
showed that the genes are classified into two subclusters, and the clustering is well correlated to the
genomic organization. This suggests that each subcluster arose independently from a separate ancestralgene. It seemed likely that members of the same subcluster of a multigene family tend to have similar expression patterns and functions. In fact, Krt1-13, -15,
and -19 are expressed in the same epithelial tissues,
such as oral cavity and esophagus, and are expected tohave similar functions. Likewise, Krt1-14 and -17 arecommonly expressed in the epithelia of hair follicle andmammary gland ducts. On the other hand, Krt1-10 and-12, which constitute another subcluster, are not expressed in the same tissues.
ACKNOWLEDGMENTS
We are grateful to Dr. Dennis Roop of Baylor College of Medicine
for his donation of the 39-noncoding region probe of the mouse
Krt1-14 gene. This study was supported by a Grant in Aid from the
Ministry of Education, Science and Culture of Japan. This paper iscontribution No. 2135 from the National Institute of Genetics, Japan.

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