TTY2: a multicopy Y-linked gene family

Abstract

Genes involved in human male sex determination and spermatogenesis are likely to be located on the Y chromosome. In an effort to identify Y-linked, testis-expressed genes, a cDNA selection library was generated by selecting testis cDNA with Y-cosmid clones. Resultant clones containing repetitive or vector material were eliminated, and 79 of the remaining clones were sequenced. Nineteen cDNAs showed homology with the TTY2 gene, and indicated that TTY2 is part of a large gene family. Screening of a panel of Y-linked cosmids revealed that the TTY2 gene family includes at least 26 members organized in 14 subfamilies. Further investigation revealed that TTY2 genes are arranged in tandemly arrayed clusters on both arms of the Y chromosome, and each gene comprises a series of tandemly arranged repeats. RT-PCR studies for two of these genes revealed that they are expressed in adult and fetal testis, as well as in the adult kidney. None of the genes investigated in detail contain an open reading frame. We conclude that the TTY2 gene family is composed of multiple copies, some of which may function as noncoding RNA transcripts and some may be pseudogenes.

Figure 1

Double fluorescent in situ hybridization (FISH) to human (a), chimpanzee (b), pigmy chimpanzee (c), and gorilla (d) metaphase chromosomes, using the cosmids 2E9 (TTY2L2A) and 2F6 (TTY2L12A) as probes. Specific signal in all species is seen only on the Y chromosome. The centromeric and banded signals are due to DAPI staining, and would appear as blue signal in a colored photograph.

Figure 2

Localization of TTY2L12A and TTY2L2A on the Y chromosome. Broken lines indicate position of YACs spanning the euchromatic and part of the heterochromatic region (Jones et al. 1994). TTY2L12A and TTY2L2A were positioned by PCR amplification of these YACs. Y chromosome intervals (as defined by Vergnaud et al. 1986; Vollrath et al. 1992) are indicated together with several Y- linked STSs (shown slightly elevated).

Figure 3

PCR amplification of tissue cDNA, using PGM1 primers (a and b) and primers specific for TTY2L12A (c) and TTY2L2A (d); the samples are as follows: 1, adult testis; 2, fetal testis; 3, adult prostate; 4, fetal brain; 5, fetal lung; 6, adult kidney; 7, fetal kidney; 8, fetal heart; 9, fetal stomach; 10, fetal intestine; 11, fetal limbs. M, DNA size marker; b_1–4, PCRs without cDNA. cDNA synthesis was carried out in presence (+) and absence (−) of reverse transcriptase.

Figure 4

Localization of the TTY2 gene and other members of this gene family on three genomic fragments (AC017019, AC023342, and AC009491); arrows indicate the relative orientation of the genes; numbers below arrows indicate the first bp of identity with the genomic fragment.

Figure 5

DOTPLOT analysis of (a) the TTY2 cDNA (3169 bp) to itself, and (b) TTY2 genomic DNA to itself; internal repeats appear as lines off the diagonal. For example in (a), the repeat of exon 7 sequence within exon 1 is clearly seen, and in (b) the tandem repeats of ∼2.4 kb are obvious. For clarity, a line drawing of the repeat elements within the gene is shown in (c).