Comparative Analysis of the Karyotypes of the Greater Long-Tailed Hamster and the Chinese Hamster

Home , Chinese hamster, Cricetulus, Mesocricetus

C 1997 The Japan Mendel Society Cytologia 62: 315-321, 1997

Kazunori Fujimoto1, Sen-ichi Oda1,*, Kazuhiro Koyasu2, Masashi Harada3 and Shin-ichi Sonta4

1 Laboratory of Animal Management, School of Agricultural Sciences, Nagoya University, Nagoya 464-01, Japan 2Department of Anatomy , School of Dentistry, Aichi-Gakuin University, Nagoya 464, Japan 3 Laboratory Animal Center , Osaka City University, Osaka 545, Japan 4 Department of Genetics , Institute for Developmental Research, Aichi Human Service Center, Kasugai 480-03, Japan

Accepted July 17, 1997

The subfamily Cricetinae (Rodentia) comprise 5 genera, including the genus Cricetulus of which comprise 11 species (Nowak and Paradiso 1983). The greater long-tailed hamster (Cricetulus trion or Tscherskia triton, abbreviated hereafter as a triton hamster) inhabits north-eastern Asia such as eastern Siberia, north-eastern China and Korea (Ellerman and Morrison-Scott 1951), and its diploid chromosome number is 28 (Tsuchiya and Won 1976). The Chinese hamster (Cricetulus griseus, 2n=22) has been successfully used as a laboratory animal, and its karyotype has been characterized by banding techniques (Ray and Mohandas 1976). In contrast, no cytogenetic analyses are available for a triton hamster. There are remarkable morphological differences between the two hamster species. The triton hamster is 5-6 times as weighty as Chinese hamster (Sonta and Semba 1980, Oda et al. 1995), and we are not able to obtain interspecific hybrid in cage. The coat color of the two hamster species also differs. That of the Chinese hamster is brown at back with black line in the center and white at belly, while that of a triton hamster is agouti at back. The triton hamster has a long tail, but Chinese hamster has a very short tail. Recently, it has been suggested that the two hamster species do not belong to the same genus Cricetulus, and the Chinese hamster is classified in Cricetulus but the triton hamster in Tscherskia (Musser and Carleton 1993). Cytogenetic data of the two hamster species may allow us the taxo- nomical approach. So, we analyzed the triton hamster karyotype based on the measurement and the banding pattern of chromosomes, in comparison with that of the Chinese hamster, and investigated the chromosome rearrangement that is related to evolutional process.

Materials and methods The triton hamster stock was derived from one pair of animals that we captured and then bred in our laboratory (Oda et al. 1995). The Chinese hamster was an inbred strain (CHS/Idr) that we have already established. Fibroblasts from the spleen, lung and tail of the two hamster species ( 5 of each species) were cultured according to the standard techniques. The karyotype was then analyzed using conventional Giemsa staining. The banding pattern analyses were performed using G-band staining according to Sumner et al. (1971), C-band staining according to Sumner (1972) and R- band staining according to Dutrillaux et al. (1973). The chromosomes of the two hamster species were measured and the banding patterns were compared.

* Correspondent author. 316 K. Fujimoto, S. Oda, K. Koyasu, M. Harada and S. Sonta Cytologia 62

Results

Karyotype analysis of the triton hamster The average size (•} S.D.) of each short arm, long arm and whole chromosome, derived from the measurement of 15 Giemsa-stained cells (Fig. 1), is shown in Table 1. Autosomes of the triton hamster consisted of 11 pairs (nos. 1-11) of acrocentric chromosomes and 2 pairs (nos. 12, 13) of

little metacentric ones. The size from chromosome no. 1 to no. 11 decreases gradually, and chromo-

some no. 12 and 13 were much smaller than the other autosomes. In sex chromosomes, the X chromosome was a middle-sized subtelocentric chromosome and the Y chromosome a small metacen-

tric chromosome. The X chromosome was smaller than chromosome no. 6 but slightly larger than chromosome no. 7, and the Y chromosome was between chromosome no. 10 and 11. A schematic

diagram, constructed by analysis of 15 G-banded and 15 R-banded cells (Figs. 2, 3), is shown in Fig. 4. Each chromosome showed a characteristic pattern. The C-banding, on the other hand, re-

vealed the presence of a large band in the centromeric region of all autosomes. In the X chromo-

1 2 3 4 5

6 8 9 10

11 12 13 XY Fig. 1. A karyotype of the greater long-tailed hamster (triton hamster, Cricetulus triton or Tscherskia triton), using conventional Giemsa staining.

Table1.Comparative size of each triton hamster chromosome (•} S.D.)

The values are calculated on the assumption that the total length of all chromosomes except Y chromosome is 100. 1997 Karyotype Analysis of the Greater Long-Tailed Hamster 317

1 2 3 4 5

6 7 8 9 10

11 12 13 X Y

Fig . 2 . A GTG-banded karyotype of the triton hamster.

1 2 3 4 5

6 7 8 9 10

11 12 13 X Y

Fig. 3 . A RBA-banded karyotype of the triton hamster. some, large dark bands in the short arm and the proximal-centromeric region of the long arm were present, and the whole Y chromosome was stained by C-banding (Fig. 5).

Comparison with Chinese hamster chromosomes R-banded chromosomes of the triton hamster (abbreviated hereafter as RT) were compared with those of the Chinese hamster (RC) (Table 2, Fig. 6). Five chromosomes of the triton hamster showed banding patterns identical to five Chinese hamster chromosomes: Chromosome no. 7 of the triton hamster (RT7) was identical to chromosome no. 5 of the Chinese hamster (RC5). Similarly, RT9, RT10, RT12 and RT13 resembled RC6, RC7, RC9 and RC10, respectively. The other six chro- 318 K. Fujimoto, S. Oda, K. Koyasu, M. Harada and S. Sonta Cytologia 62

1 2 3 4 5 6 7

8 9 10 11 12 13 X Y

Fig. 4 . Ideogram of GTG- and RBA-banding pattern of the triton hamster. Each chromosome is identi- fied by GTG-banding pattern.

Fig . 5 . A C-banded karyotype of the triton hamster. mosomes of the triton hamster were homologous to the respective arms of three Chinese hamster chromosomes. RT1 was similar to the long arm of RC1. Similarly, RT3, RT6, RT5, RT4 and RT8 corresponded respectively to the short arm of RC1, the long arm of RC2, the short arm of RC2, the long arm of RC3 and the short arm of RC3. When we consider inversion at a certain region of chromosomes, the remaining two chromosomes of the triton hamster of RT2 and RT11, were similar to Chinese hamster chromosomes of RC4 and RC8, respectively. In sex chromosomes, the short arm of RCX was homologous to the proximal-telomeric region of the long arm of RTX, and a part of 1997 Karyotype Analysis of the Greater Long-Tailed Hamster 319

Table2.Chromosomal comparison of the Triton hamster the long arm of RCX also corresponded to the with the Chinese hamster RTX. RCY was almost similar to RTY. While large C-bands existed near centromeric region in most autosomes of the triton hamster, small C-bands near centromeric region in most autosomes and bands in intermediate part of both arms of no. 1 were present in the Chinese hamster.

Discussion While Musser and Carleton (1993) suggested recently that the triton hamster and the Chinese hamster do not belong to the same genus Cricetulus, others also reported that they did (Nowak and Paradiso 1983). Nevertheless, there is a significant difference in the diploid * inv.=pericentric inversion. chromosome number and chromosome mor- ** Xp of the Chinese hamster is homologous to Xq of the triton hamster, but no region homologous to the distal phology between them. However, the banding region of Xq of the Chinese hamster existed in the triton patterns showed clearly the homology of chro- hamster chromosome. mosomes; each chromosome of the triton ham- *** The centromeric region of Y of the Chinese hamster ster corresponded to one arm or one whole is similar to that of the triton hamster, but no segment of Y of the triton hamster corresponding to the distal region of chromosome of the Chinese hamster. Generally Yq of the Chinese hamster. speaking, morphological changes such as peri-

T8 T5 TX TY T1 C1 C2 C3 C4 T2 CX CY T6 T4

C5 T7 C6 T9 C7 T10 C8 T11 C9 T12 C10T13

Fig. 6. Comparison of RBA-banded chromosomes of the triton hamster (T) with those of the Chinese hamster (C). The banding pattern of each triton hamster chromosome is similar to that of an arm or a chromosome of the Chinese hamster. On the assumption of T2 and T11 is similar to that of C4 and C8, respectively. 320 K. Fujimoto, S. Oda, K. Koyasu, M. Harada and S. Sonta Cytologia 62 centric inversions, Robertsonian fusions or centromeric fissions play an important role in the process of karyotypical evolution (Fredga 1977, Holmquist and Dancis 1980, Schmid et al. 1986). Thus, the correspondence of chromosomes between two hamster species may serve as a plausible explanation for the karyotypical changes accompanying evolution. On the other hand, there were distinctly C-bands around the centromeric region in the triton hamster chromosomes but such centromeric bands were small or none in Chinese hamster chromosomes. Data from G- and R-banding indicated that chromosome no. 1 of the Chinese hamster, for example, corresponded to a figure assumed by Robertsonian fusion between chromosome nos. 1 and 3 of the triton hamster, or the reverse case by fission. However, there is no C-band in the centromeric region but pale bands in both intermediate regions of the short arm (1p26) and the long arm (1q17). On the assumption that the common ancestor animal of both the triton hamster and Chinese hamster had the same karyotype as the present triton hamster, the heterochromatic region around the centromere of chromosome nos. 1 and 3 in the ancestor animal might be transposed to the intermediate region of each arm by insertion or inversion including these regions. Furthermore, a part of the heterochromatic bands might have been lost accompanied with or without such rearrangements. Similar changes of size of heterochromatic bands might occur accompanied with or without the occurrence of Robertsonian fusions in many chromosomes of the ancestor animal. Cer- tainly, one can assume the reverse case in which the common ancestor animal had a karyotype similar to that of the present Chinese hamster, and the mechanism of karyological changes might be centromeric fission from bi-arm chromosomes. The morphological differences of the X chromosome between the two hamster species, such as size and position of the centromere, might be also caused by rearrangements of the X including heterochromatic region. The significant differences in the C-band regions among hamster species are interesting in studies on karyotypical evolution among species and genera. The biological meaning of constitutive heterochromatin remains unclear. Moreover, none had determined the association of increase and decrease of heterochromatin evolution of species. Among karyotypes of some species belonging to a closely related genera, a significant difference in the size of C-bands is often seen (Murray et al. 1980, Popescu and DiPaolo 1980, Stock 1981). Similarly, individuals in the same species as well as those among subspecies often show polymorphism of C-bands (Forejt 1973, Arnason et al. 1980). The increase and decrease of such heterochromatin usually has no influence on the phenotype. These findings suggest that the amount of heterochromatin in the genome changes without evolution of species. The volume and position of heterochromatin in one species might have been fixed whether a dominant population had more heterochromatin or less heterochromatin during speciation. Although the triton hamster chromosomes appeared to be homologous to the Chinese hamster ones, further studies such as comparative analysis using flourescent in situ hybridization (FISH) and analysis at the DNA level are needed to determine the genome homology between the two hamster species. It is difficult to say which the triton hamster is belonging to the same genus Cricetulus or different genus Tscherskia. We need more informations of the biological data from the triton hamster and the related species.

Summary The G-, R- and C-banded karyotypes of the two hamster species, the greater long-tailed hamster (Cricetulus triton or Tscherskia triton) and the Chinese hamster (Cricetulus griseus), were analyzed. The diploid chromosome number of Chinese hamster is 22, and that of the greater long- tailed hamster 28. These are morphological differences between two hamster species: the number of metacentric and submetacentric autosomes was only 4 in the greater long-tailed hamster, in contrast to 14 in the Chinese hamster, while the number of acrocentric and subtelocentric chromosome was 1997 Karyotype Analysis of the Greater Long-Tailed Hamster 321

22 in the greater long-tailed hamster and 6 in the Chinese hamster. The analysis of banding patterns in the two hamster species revealed that each chromosome of the greater long-tailed hamster corresponded to one arm or one whole chromosome of the Chinese hamster. However, the large C-bands (or lightly stained with R-bands) such as those found around the centromeric region of the greater long-tailed hamster chromosomes did not exhist in the centromeric region, or any other segments, of any chromosome in the Chinese hamster. These findings may be related to the karyotypical evolution of the two hamster species, but it is difficult to say which these hamster species are belonged to the same genus Cricetulus or not.

Key words : Karyotype, Cricetulus triton, Tscherskia triton, Cricetulus griseus, Banding pattern

Acknowledgement A part of this study was supported by a grant-in-aid for scientific research of the Ministry of Education, Science, Sports and Culture (No. 0750806) and the Tutikawa Memorial Fund for Study in Mammalian Mutagenecity.

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