Cytogenetic and Genome Research

Banga, S.S.; Peng, L.; Dasgupta, T.; Palejwala, V.; Ozer, H.L.

doi: 10.1159/000251960pmid: 20068294

Normal human diploid fibroblasts have limited life span in culture and undergo replicative senescence after 50–60 population doublings. On the contrary, cancer cells typically divide indefinitely and are immortal. Expression of SV40 large T and small t antigens in human fibroblasts transiently extends their life span by 20–30 population doublings and facilitates immortalization. We have identified a rearrangement in chromosome 6 shared by SV40-transformed human fibroblasts. Rearrangements involving chromosome 6 are among the most frequent in human carcinogenesis. In this paper, we extend analysis of the 6q26–q27 region, a putative site for a growth suppressor gene designated SEN6 involved in immortalization of SV40-transformed cells. Detailed molecular characterization of the rearranged chromosomes (6q*, normal appearing; and 6q^t, translocated) in the SV40-immortalized cell line HALneo by isolating each of these 2 chromosomes in mouse/HAL somatic cell hybrids is presented. Analysis of these mouse/HAL somatic cell hybrids with polymorphic and nonpolymorphic markers revealed that the 6q* has undergone a chromosomal break in the MLLT4 gene (alias AF6). This result in conjunction with previous published observations leads us to conclude that SEN6 lies between MLLT4 and TBP at chromosomal region 6q27. Examination of different genes (MLLT4, DLL1, FAM120B, PHF10) located within this interval that are expressed in HS74 normal fibroblast cells reveals that overexpression of epitope-tagged truncated PHF10 cDNAs resulted in reduced cell proliferation in multiple cell lines. Paradoxically, down-regulation of PHF10 by RNAi also resulted in loss of cell proliferation in normal fibroblast cells, indicating PHF10 function is required for cell growth. Taken together, these observations suggest that decreased cell proliferation with epitope-tagged truncated PHF10 proteins may be due to dominant negative effects or due to unregulated expression of these mutant proteins. Hence we conclude that PHF10 is not SEN6 but is required for cell growth.

Kaczmarek, K.; Niedzialkowska, E.; Studencka, M.; Schulz, Y.; Grzmil, P.

doi: 10.1159/000251961pmid: 20068295

Many genes crucial for male fertility are often predominantly or exclusively expressed in male germ cells. The analysis of mouse models has demonstrated the functional importance of peroxisomes in spermatogenesis. The CCDC33 protein has been reported to be a cancer/testis (CT) antigen. We found that mouse Ccdc33 is predominantly expressed in the testis and undergoes alternative splicing to produce at least 4 different transcripts. The protein encoded by Ccdc33 contains 3 coiled-coil domains, a C2-domain, 2 ER membrane retention signal-like motifs and 2 putative peroxisomal targeting signals type 2 (PTS2). We could demonstrate that the second PTS2 sequence is functional and responsible for the targeting of CCDC33 to peroxisomes. Moreover, in HeLa cells CCDC33-dsRED fusion protein co-localized with a known peroxisomal protein, namely PXT1, and showed punctuate intracellular distribution. Taken together, the mouse Ccdc33 encodes a putative peroxisomal protein and is predominantly expressed in male germ cells. The expression starts at the primary spermatocyte stage, suggesting an important role of this protein during spermatogenesis.

Pita, M.; Gosálvez, J.; Gosálvez, A.; Nieddu, M.; López-Fernández, C.; Mezzanotte, R.

doi: 10.1159/000251962pmid: 20068296

Significant similarity between human and gorilla genomes has been found in all chromosome arms, but not in centromeres, using whole-comparative genomic hybridization (W-CGH). In human chromosomes, centromeric regions, generally containing highly repetitive DNAs, are characterized by the presence of specific human DNA sequences and an absence of homology with gorilla DNA sequences. The only exception is the pericentromeric area of human chromosome 9, which, in addition to a large block of human DNA, also contains a region of homology with gorilla DNA sequences; the localization of these sequences coincides with that of human satellite III. Since highly repetitive DNAs are known for their high mutation frequency, we hypothesized that the chromosome 9 pericentromeric DNA conserved in human chromosomes and deriving from the gorilla genome may thus play some important functional role.

Chiba, M.; Kiyosawa, H.; Hiraiwa, N.; Ohkohchi, N.; Yasue, H.

doi: 10.1159/000251963pmid: 20068297

PTEN-induced kinase 1 (PINK1), which is identified as the gene transactivated by the tumor suppressor PTEN, has been found to be one of the causative genes in Parkinson’s disease (PD). In order to understand PD, rodent models containing affected Pink1 such as loss-of-function mutations have been exploited. Recently, natural antisense RNA of PINK1 has been demonstrated to be involved in the regulation of the PINK1 locus. However, no antisense RNAs of Pink1 except for human have been reported so far. Therefore, in the present study, while searching for the Pink1 antisense RNAs in mouse, we found that the antisense RNAs are transcribed from a mouse genomic region corresponding to the human region from which the antisense RNAs are produced. Further, we investigated the localization of the antisense RNAs in mouse brain using in situ hybridization; this demonstrated that the antisense RNAs were localized in the regions of brain where the Pink1 mRNA was found. In addition, the mRNA and antisense RNAs were found more densely in the hippocampus than in the other brain regions in newborn and 1-week-old mice, while those RNAs were found uniformly in the mouse brain regions of embryo day (E) 14, E17, and 8-weeks-old.

Quach, T.A.; Villagómez, D.A.F.; Coppola, G.; Pinton, A.; Hart, E.J.; Reyes, E.R.; Basrur, P.K.; King, W.A.

doi: 10.1159/000251964pmid: 20068298

Chromosome abnormalities are well known for their negative impact on the reproductive performance of carriers. Such abnormalities could have severe effect on animal industries which rely heavily on efficient reproduction. We conducted a cytogenetic survey of breeder pigs from 4 different Canadian farms to investigate the frequency of chromosome abnormalities and to assess their reproductive impact on pig populations. Our study revealed that 50% of the ‘hypoprolific’ boars and 2.5% of the young boars raised for service in artificial insemination were carriers of chromosome anomalies while no chromosome defect was noted in any of the ‘proven’ breeder boars. G-banding technique to determine the type of abnormalities detected 3 previously unreported translocations involving chromosomes 1 and 6, chromosomes 10 and 13 and chromosomes 9 and 14. The reciprocal nature of these translocations was confirmed either using fluorescent in situ hybridization (FISH) technique or immunostaining for synaptonemal complex delineation and were named rcp(1;6)(p22,q12), rcp(10;13), and rcp(9;14) (p24;q27), respectively. Prolificacy of 1/6 and 10/13 translocation carriers was noted to be reduced by more than 40% compared to their normal counterparts while it was reduced by 26% in carriers of the 9/14 translocation. Carriers of 1/6 and 9/14 translocations displayed a higher repeat breeding tendency, compared to their herd average (5 and 16%, respectively). While for the 9/14 translocation the prevalence of stillbirths was lower than that in their herd [8.7 vs. 10.4% (p < 0.001)]. The present results, albeit based on a relatively small number of pigs, indicate that the prevalence of chromosome abnormalities could be much higher in Canadian pigs compared to that reported in European pigs and underline the urgent need to initiate cytogenetic screening programs as one of the effective ways to reduce reproductive problems in Canadian pig populations.

Hansmann, T.; Nanda, I.; Volobouev, V.; Yang, F.; Schartl, M.; Haaf, T.; Schmid, M.

doi: 10.1159/000251965pmid: 20068299

The stone curlew, also known as thick-knee (Burhinus oedicnemus, BOE), represents a phylogenetically young species of the shorebirds (Charadriiformes) that exhibits one of the most atypical genome organizations known within the class of Aves, due to an extremely low diploid number (2n = 42) and only 6 pairs of microchromosomes in its complement. This distinct deviation from the ‘typical’ avian karyotype is attributed to repeated fusions of ancestral microchromosomes. In order to compare different species with this atypical avian karyotype and to investigate the chromosome rearrangement patterns, chromosome-specific painting probes representing the whole genome of the stone curlew were used to delineate chromosome homology between BOE and 5 species belonging to 5 different avian orders: herring gull (Charadriiformes), cockatiel (Psittaciformes), rock pigeon (Columbiformes), great gray owl (Strigiformes) and Eurasian coot (Gruiformes). Paints derived from the 20 BOE autosomes delimited 28 to 33 evolutionarily conserved segments in the karyotypes of the 5 species, similar to the number recognized by BOE paints in such a basal lineage as the chicken (28 conserved segments). This suggests a high degree of conservation in genome organization in birds. BOE paints also revealed some species-specific rearrangements. In particular, chromosomes BOE1–4 and 14, as well as to a large extent BOE5 and 6, showed conserved synteny with macrochromosomes, whereas homologous regions for BOE7–13 are found to be largely distributed on microchromosomes in the species investigated. Interestingly, the 6 pairs of BOE microchromosomes 15–20 appear to have undergone very few rearrangements in the 5 lineages investigated. Although the arrangements of BOE homologous segments on some chromosomes can be explained by complex fusions and inversions, the occurrence of homologous regions at multiple sites may point to fission of ancestral chromosomes in the karyotypes of the species investigated. However, the present results demonstrate that the ancestral microchromosomes most likely experienced fusion in the stone curlew lineage forming the medium-sized BOE chromosomes, while they have been conserved as microchromosomes in the other neoavian lineages.

Hulick, P.J.; Noonan, K.M.; Kulkarni, S.; Donovan, D.J.; Listewnik, M.; Ihm, C.; Stoler, J.M.; Weremowicz, S.

doi: 10.1159/000251966pmid: 20068300

Approximately 15 patients with partial trisomy 9p involving de novo duplications have been previously described. Here, we present clinical, cytogenetic, FISH and aCGH findings in a patient with a de novo complex rearrangement in the short arm of chromosome 9 involving an inverted duplication at 9p24→p21.3 and a deletion at 9pter→p24.2. FISH probes generated from BACs selected from the UCSC genome browser were utilized to verify this rearrangement. It is likely that some previously described duplications of 9p may also be products of complex chromosomal aberrations. This report in which FISH and aCGH were used to more comprehensively characterize the genomic rearrangement in a patient with clinical manifestations of 9p duplication syndrome underscores the importance of further characterizing cytogenetically detected rearrangements.