Using chromosomal in situ hybridization it has been demonstrated that specific members of the YRRM and the TSPY families are multicopy and Y chromosome specific in hominoids. After hybridization with the YRRM-related cosmid A5F and the TSPY-related cosmids cos36 and cY91, a reverse and complementary pattern of main and secondary signals is detected on the Y chromosomes of the human, the pygmy chimpanzee and the gorilla, while the location of signals coincides on the Y chromosomes of the chimpanzee, both orang-utan subspecies and the white hand gibbon. This complementary distribution of YRRM and TSPY sequences on the hominoid Y chromosomes possibly originates from a similar sequence motif that is shared by and evolutionarily conserved between certain members of both gene families and/or repeated elements flanking those genes. Otherwise this complementary distribution could go back to a common organization of these genes next to each other on an ancient Y chromosome which was disrupted by chromosomal rearrangements and amplification of one or other of the genes at each of the locations.

The three human male specific expressed gene families DAZ, RBM, and TSPY are known to be repetitively clustered in the Y-specific region of the human Y Chromosome (Chr). RBM and TSPY are Y-specifically conserved in simians, whereas DAZ cannot be detected on the Y chromosomes of New World monkeys. The proximity of SRY to the pseudoautosomal region (PAR) is highly conserved and thus most effectively stabilizes the pseudoautosomal boundary on the Y (PABY) in simians. In contrast, the non-recombining part of the Y Chrs, including DAZ, RBM, and TSPY, was exposed to species-specific amplifications, diversifications, and rearrangements. Evolutionary fast fixation of any of these variations was possible as long as they did not interfere with male fertility.

Cytogenetic studies of the primate Y chromosomes have suggested that extensive rearrangements have occurred during evolution of the great apes. We have used in situ hybridization to define these rearrangements at the molecular level. pHU-14, a probe including sequences from the sex determining gene SRY, hybridizes close to the early replicating pseudoautosomal segment in a telomeric or subtelomeric position of the Y chromosomes of all great apes. The low copy repeat detected by the probe Fr35-II is obviously included in Y chromosomal rearrangements during hominid evolution. These results, combined with previous studies, suggest that the Y chromosome in great apes has a conserved region including the pseudoautosomal region and the testis-determining region. The rest of the Y chromosome has undergone several rearrangements in the different great apes.

Applying two-colour fluorescence in situ hybridization (FISH) we simultaneously hybridized RBM- and TSPY-related cosmids to Y chromosomes in prophase and to released Y chromatin in interphase nuclei of man and pygmy chimpanzee. Whereas, even on prophasic Y chromosomes, no resolution of the overlapping RBM and TSPY signal clusters could be achieved, the RBM and TSPY signals are completely separated from each other in our maximum released Y chromatin stretches in interphase nuclei. These results unequivocally lend support to the view that the RBM and TSPY families have an interspersed organization on the Y chromosomes of man and higher apes. Thus, the distribution of RBM and TSPY signals might well go back to a common organization of these genes next to each other on an ancient Y chromosome.

Twenty-five specimens of lowland gorilla, including 24 specimens of the western lowland gorilla (Gorilla gorilla gorilla) and 1 specimen of the eastern lowland gorilla (G. gorilla graueri), were investigated by fluorescence in situ hybridization with a human-derived 18S + 28S rDNA probe. Specific hybridization was constitutively seen on the short arms of gorilla acrocentric chromosome pairs 22 and 23, corresponding to human pairs 21 and 22. Only one specimen of western lowland gorilla investigated showed an additional hybridization site at the telomeric short arm of one chromosome 1. From our own results and those in the literature, it is clear that the additional rDNA site on chromosome 1 must be regarded as a rare polymorphism in the subspecies of western lowland gorilla, possibly going back to a founder translocation event.