Arms Of Acrocentric Chromosomes Research Articles

STRavinsky STR database and PGTailor PGT tool demonstrate superiority of CHM13-T2T over hg38 and hg19 for STR-based applications.

Short-Tandem-Repeats (STRs) have long been studied for possible roles in biological phenomena, and are utilized in multiple applications such as forensics, evolutionary studies and pre-implantation-genetic-testing (PGT). The two reference genomes most used by clinicians and researchers are GRCh37/hg19 and GRCh38/hg38, both constructed using mainly short-read-sequencing (SRS) in which all-STR-containing-reads cannot be assembled to the reference genome. With the introduction of long-read-sequencing (LRS) methods and the generation of the CHM13 reference genome, also known as T2T, many previously unmapped STRs were finally localized within the human genome. We generated STRavinsky, a compact STR database for three reference genomes, including T2T. We proceeded to demonstrate the advantages of T2T over hg19 and hg38, identifying nearly double the number of STRs throughout all chromosomes. Through STRavinsky, providing a resolution down to a specific genomic coordinate, we demonstrated extreme propensity of TGGAA repeats in p arms of acrocentric chromosomes, substantially corroborating early molecular studies suggesting a possible role in formation of Robertsonian translocations. Moreover, we delineated unique propensity of TGGAA repeats specifically in chromosome 16q11.2 and in 9q12. Finally, we harness the superior capabilities of T2T and STRavinsky to generate PGTailor, a novel web application dramatically facilitating design of STR-based PGT tests in mere minutes.

31. Rare familial rearrangement with acrocentric satellite material on Yp relocating to Xp in a female fetus

The short (p) arms of acrocentric chromosomes contain chromatin material described as stalks and satellites made up of repetitive DNA sequences including those encoding ribosomal RNA, and correspond to the nucleolar organizer regions (NOR). Satellites observed on non-acrocentric chromosomes are rare aberrations, typically resulting from translocation between the p-arm of an acrocentric chromosome and the distal arm of another chromosome. The phenotypic consequences of such unbalanced rearrangements can vary significantly depending on the size and gene content of the non-acrocentric chromosome involved.

TBP-Related Factor 2 as a Trigger for Robertsonian Translocations and Speciation.

Robertsonian (centric-fusion) translocation is the form of chromosomal translocation in which two long arms of acrocentric chromosomes are fused to form one metacentric. These translocations reduce the number of chromosomes while preserving existing genes and are considered to contribute to speciation. We asked whether hypomorphic mutations in genes that disrupt the formation of pericentromeric regions could lead to centric fusion. TBP-related factor 2 (Trf2) encodes an alternative general transcription factor. A decrease of TRF2 expression disrupts the structure of the pericentromeric regions and prevents their association into chromocenter. We revealed several centric fusions in two lines of Drosophila melanogaster with weak Trf2 alleles in genetic experiments. We performed an RNAi-mediated knock-down of Trf2 in Drosophila and S2 cells and demonstrated that Trf2 upregulates expression of D1—one of the major genes responsible for chromocenter formation and nuclear integrity in Drosophila. Our data, for the first time, indicate that Trf2 may be involved in transcription program responsible for structuring of pericentromeric regions and may contribute to new karyotypes formation in particular by promoting centric fusion. Insight into the molecular mechanisms of Trf2 function and its new targets in different tissues will contribute to our understanding of its phenomenon.

Jumping translocations of chromosome 1q occurring by a multi-stage process in an acute myeloid leukemia progressed from myelodysplastic syndrome with a TET2 mutation

BackgroundJumping translocations (JTs) are rare chromosome rearrangements characterized by re-localization of one donor chromosome to multiple recipient chromosomes. Here, we describe an acute myeloid leukemia (AML) that progressed from myelodysplastic syndrome (MDS) in association with acquisition of 1q JTs. The sequence of molecular and cytogenetic changes in our patient may provide a mechanistic model for the generation of JTs in leukemia.Case presentationA 68-year-old man presented with pancytopenia. Bone marrow aspirate and biopsy showed a hypercellular marrow with multilineage dysplasia, consistent with MDS, with no increase in blasts. Karyotype and MDS fluorescence in situ hybridization (FISH) panel were normal. Repeat bone marrow aspirate and biopsy after 8 cycles of azacitidine, with persistent pancytopenia, showed no changes in morphology, and karyotype was again normal. Myeloid mutation panel showed mutations in RUNX1, SRSF2, ASXL1, and TET2. Three years after diagnosis, he developed AML with myelodysplasia-related changes. Karyotype was abnormal, with unbalanced 1q JTs to the short arms of acrocentric chromosomes 14 and 21, leading to gain of 1q.ConclusionsOur patient had MDS with pathogenic mutations of the RUNX1, SRSF2, ASXL1, and TET2 genes and developed 1q JTs at the time of progression from MDS to AML. Our data suggest that the formation of 1q JTs involves multiple stages and may provide a mechanistic model for the generation of JTs in leukemia.

70. PREIMPLANTATION GENETIC TESTING FOR YQH-AUTOSOME TRANSLOCATION CARRIERS: ANALYSIS OF GAMETE SEGREGATION

Introduction Y-autosome translocations occur in approximately 1:2000 in population and represent special group of reciprocal exchanges. Due to a balanced nature they do not influence on phenotype in general. However, Y-autosome translocation can dislocate sex bivalent formation and chromosome synapsis in the first meiotic division prophase that led to gamete aneuploidy or pachytene arrest. More often Y-derived DNA, predominantly Yq12 band, is translocated onto the short arm of acrocentric chromosomes, mainly chromosomes 15 and 22 due to a strong correlation in their satellite DNA sequences. In women inheritance of such derivative acrocentric chromosomes could be associated with the risk of ovarian malignancy, but the particular mechanism of this pathological effect is not entirely clear since the Yq12 band consists of tandemly organized satellite DNA sequences. It can be associated with the transcription of satellite DNAwhich occurs during carcinogenesis. Material & methods Two families with Yq12/15p and one – with Yq12/22p translocations were analyzed during IVF cycle with PGT in International Centre for Reproductive Medicine. The mean age of men and women was 36±2.8 and 34.25±1.4 years respectively. Karyotyping analysis was performed using GTG banding technique on metaphase chromosomes from peripheral blood lymphocytes. Fluorescence in situ hybridization (FISH) was used to clarify the breakpoint on chromosomes and confirm the chromosomes-specificity of the probes. Additionally, seven chromosomes (13, 14, 15 or 22, 16, 18, 21, X), which are not involved in rearrangement, were analyzed. Results FISH with chromosome specific probes was performed on blastomere from cleavage-stage embryo. A total of 42 blastomeres were analyzed. In 43% of the blastomeres «adjacent-1» segregationwith derivate autosome and X or Y chromosomes was revealed. Segregation 3:1 was observed in 10% of the cells, with no predominant inheritance of the derivative autosome. In 21% of cells the type of chromosome segregation could not be determined due to mosaicismor polyploidy. 45% of the cells were aneuploid for autosomes, which are not involved in the translocation, which may be connected with interchromosomal effect. Conclusions Carriers of Yq12/15p or Yq12/22p translocations have a predominant inheritance of the derivative autosome, which can be associated with the disruption ofsex bivalent segregation. This research was supported by Russian Foundation for Basic Research (grant # 18-34-00279 ).

Cytogenetic and phylogenetic studies of diploid and polyploid members of Tribe Anemoninae (Ranunculaceae)

The ancestry, phylogenetic differentiation and systematic classification of the worldwide-distributed genus Anemone have been debated for many years. In this paper 11 Anemone, three Pulsatilla species and Hepatica nobilis were subjected to detailed karyotype analysis with the aim of obtaining new cytogenetic data that will contribute to karyotype evolutionary studies of the tribe Anemoninae. The results are interpreted in a phylogenetic context, established from the intergenic nontranscribed spacer (NTS) of 5S rDNA and internal transcribed spacer (ITS) of 35S rDNA. One to three 35S and one to three 5S rDNA loci are present in diploid and polyploid taxa. The 35S rDNA loci are located terminally on the short arm of acrocentric chromosomes, while for 5S rDNA there is no preferential chromosomal position as it exhibits terminal, subterminal, interstitial or pericentromeric positions, and is located either on acrocentric or metacentric chromosomes. The karyotype of hexaploid A. baldensis (2n = 6x = 48) is presented for the first time, and A. sylvestris is proposed as one of its putative parental species. Chromosome fusion/translocation is proposed as the key mechanism involved in reduction of the basic chromosome number from 8 in the Anemone subgenus to 7 in the Anemonidium subgenus. The cytogenetic data obtained are mainly supported by ITS and NTS phylogeny. Diversification of the genus Anemone was accompanied by a large reduction of heterochromatin, from the Mediterranean anemones that have large amounts of heterochromatin to the New World anemones without any detectable heterochromatic blocks.

The boundary of macaque rDNA is constituted by low-copy sequences conserved during evolution

In Macaca mulatta, the single rDNA array is flanked by a patchwork of sequences including subregions of human Yp11.2, 4q35.2, and 10p15.3. This composite DNA region is characterized by unique or low-copy sequences, resembling a potentially transcribed region. The analysis of Cercopithecus aethiops, Presbytis cristata, and Hylobates lar suggests that this complex sequence organization could be shared by Old World monkey and lesser ape species. After the lesser apes/great apes divergence, the unique or nonduplicated DNA region underwent amplification and spreading, preferentially marking the p arm of acrocentric chromosomes bearing the rDNA. The molecular analysis of human acrocentric chromosomes revealed some extent of remodeling of the rDNA boundary: near the human NOR, a large 4q35.2 duplication partially resembles that found in MMU; conversely, infrequently represented Yp11.2 sequences totally differed from those of the macaque, and 10p15.3 sequences were lacking. Thus, although evolutionary events modified the sequence organization of the MMU rDNA boundary, its overall sequence feature and the preferential location in vicinity to the NOR have been conserved.

Segment duplications in subtelomeric regions of human chromosome 13

Owing to a great progress in studying the human genome, its euchromatic portion is almost completely sequenced; the complete sequence is still unknown only for pericentric and telomeric regions and short arms of acrocentric chromosomes. Extended satellite blocks and segment duplications located in these regions substantially hinder the joining of the sequenced fragments and construction of the full-length genome map. The sequence was established for a 1.5-kb human chromosome 13 subtelomeric region, which is about 10 kb away from the rDNA cluster, and deposited in GenBank under accession no. AF478540. The region showed 83-84% homology to the pericentric region of human chromosome 19, and contained short fragments homologous to the pericentric region of human chromosome 13. The results may contribute to the current revision of genome evolution concepts in view of numerous segment duplications revealed.

First Chromosome Data about the silverside Atherinella brasiliensis (Atheriniformes, Pisces) from the South coast of Brazil. Conventional, C- NOR and CMA3 bandings and FISH studies

Abstract Cytogenetic studies of A. brasiliensis from the south coast of Brazil were made with specimens of two populations: Baia de Paranagua and Lagoa da Conceicao. All the individuals analyzed presented a diploid number of 48 chromosomes with a standard karyotype constituted by 4M + 10SM + 14ST + 20A (FN=76). There have been found five different cytotypes (A to E) in the specimens of the Baia de Paranagua/PR, while in the Lagoa da Conceicao/SC just two (standard and F). These cytotypes are characterized by changes in the number of M and SM chromosomes and these differences could be due to pericentric inversions. The C-banding regions were evidenced in pericentromeric and telomeric regions also staining positively the secondary constrictions. NORs detected by AgNO3 and CMA3 staining are located at the short arms of acrocentric chromosomes (19th pair) and these techniques evidence a size heteromorphism between the homologue chromosomes. The heteromorphism of the NOR bearing chromosome was not detect by FI...

Human genome dispersal and evolution of 4q35 duplications and interspersed LSau repeats

We have investigated the evolutionary history of the 4q35 paralogous region, and of a sub-family of interspersed LSau repeats. In HSA, 4q35 duplications were localized at 1q12, 3p12.3, 4q35, 10q26, 20cen, whereas duplicons and interspersed LSau repeats simultaneously labeled the p arm of acrocentric chromosomes. A multi-site localization of 4q35-like sequences was also observed in PTR, GGO, PPY, HLA (Hominoidea) and PAN (Old World monkey), thus indicating that duplications of this region have occurred extensively in the two clades, which diverged at least 25 million years ago. In HSA, PTR and PAN, 4q35-derived duplicons co-localized with rDNA, whereas in GGO and PPY this association was partially lacking. In PAN, the single- and multi-site distribution of rDNA and paralogous sequences, respectively, indicates a different timing of sequence dispersal. The sub-family of interspersed LSau repeats showed a lesser dispersal than 4q35 duplications both in man and great apes. This finding suggests that duplications and repeated sequences have undergone different expansion/contraction events during evolution. The mechanisms underlying the dispersal of paralogous regions may be further derived through studies comparing the detailed structural organization of these genomic regions in man and primates.

Human acrocentric chromosomes with transcriptionally silent nucleolar organizer regions associate with nucleoli.

Human ribosomal gene repeats are distributed among five nucleolar organizer regions (NORs) on the p arms of acrocentric chromosomes. On exit from mitosis, nucleoli form around individual active NORs. As cells progress through the cycle, these mini-nucleoli fuse to form large nucleoli incorporating multiple NORs. It is generally assumed that nucleolar incorporation of individual NORs is dependent on ribosomal gene transcription. To test this assumption, we determined the nuclear location of individual human acrocentric chromosomes, and their associated NORs, in mouse> human cell hybrids. Human ribosomal genes are transcriptionally silent in this context. Combined immunofluorescence and in situ hybridization (immuno-FISH) on three-dimensional preserved nuclei showed that human acrocentric chromosomes associate with hybrid cell nucleoli. Analysis of purified nucleoli demonstrated that human and mouse NORs are equally likely to be within a hybrid cell nucleolus. This is supported further by the observation that murine upstream binding factor can associate with human NORs. Incorporation of silent NORs into mature nucleoli raises interesting issues concerning the maintenance of the activity status of individual NORs.

The structure of duplications on human acrocentric chromosome short arms derived by the analysis of 15p

We report the molecular analysis of a 130-kb DNA region containing a junction between beta and non-beta satellite DNA from chromosome 15p. The genomic region is characterized by beta satellite blocks intermingled with variants of the D4Z4 repeat, and duplicons from 4q24 and 4q35. Besides the p-arm of acrocentric chromosomes, the duplicons showed a wide genomespread involving pericentromeric, sub-telomeric, and interstitial regions. In this regard, the paralogous sequences were characterized by a high similarity index (96%), thus indicating a recent transposition during the evolution. The acrocentrics differedwith regard to the location of the 4q24 paralogous region, since it mapped on the p-arm of chromosomes 13-15 and 21, but only on 22q11.2. Conversely, the 4q35 duplication marked the p-arm of all the acrocentrics. In different individuals, the short arm of acrocentric chromosomes revealed a great variability of sequence representation and location at p11 and/or p13 for both the 4q24 and 4q35 duplications. The studied genomic region from chromosome 15p, of which a contig of approximately 200 kb has been derived, could lead to more detailed investigations into the sequence organization and possible biological function of chromosome regions that are located close to the rDNA array.

PRINS analysis of the telomeric sequence in seven lemurs.

We examined the chromosomal localization of the telomeric sequence, (TTAGGG)n, in seven species of the lemurs and one greater galago, as an outgroup, using the primed in-situ labeling (PRINS) technique. As expected, the telomeric sequence was identified at both ends of all chromosomes of the eight prosimians. However, six species showed a signal at some pericentromeric regions involving constitutive heterochromatin as well. The pericentromeric region of chromosome 1 of Verreaux's sifaka (Propithecus verreauxi verreauxi) was labeled with a large and intense signal. The range of the signal considerably exceeded the area of DAPI positive heterochromatin. On the other hand, in the five lemurs, a large signal was detected also in the short arm of acrocentric chromosomes. Acquisition of the large block of the telomeric sequence into the acrocentric short arm might be interpretable in terms of the tandem growth of the heterochromatic short arm and the reciprocal translocation between heterochromatic short arms involving the telomeric sequence. Subsequently, it was postulated that meta- or submetacentric chromosomes possessing the telomeric sequence at the pericentromeric region could be formed by centric fusion between such acrocentric chromosomes.

AgNORs in Benign, Dysplastic, and Malignant Melanocytic Skin Lesions

Nucleolar organizer regions (NORs) are loops of DNA situated on the short arms of acrocentric chromosomes 13, 14, 15, 21, and 22; they can be demonstrated in formalin-fixed paraffin-embedded sections by a one-step silver technique, the resulting black structures being termed AgNORs. We have applied the technique to 30 benign "banal" nevi (BN), 30 dysplastic nevi (DN), and 30 malignant melanomas (MM). AgNORs in 200 nuclei were scored and the means calculated. Counts were as follows: BN showed a mean of 1.3 AgNORs per nucleus within a range of 1.1-1.6; DN showed a mean of 1.2 within a range of 1.0-1.6; and MM showed a mean of 2.1 within a range of 1.2-4.2. A significant difference existed between counts for MM and those for BN and DN, despite some overlap. There was no statistically significant difference between BN and DN. Although still within the field of research, the AgNOR technique may prove to be of value in helping to differentiate MM from DN, but is unlikely to be of help in separating DN from BN.

Molecular cytogenetic evidence to characterize breakpoint regions in Robertsonian translocations

Four individuals carrying different Robertsonian translocations (13q;14q, 14q;21q, 14q;15q, and 13q;21q) were studied to determine the breakpoints involved in the generation of these derivative chromosomes. Sequential high-resolution G-banding, in situ hybridization using alphoid and ribosomal DNA probes, and C-banding were performed. In addition, silver staining was also used for visualization of the NOR region. The results provide direct molecular cytogenetic evidence that Robertsonian translocations can take place in different regions in both the short arm and proximal long arm of acrocentric chromosomes. Three different types of breakpoints were identified: between the ribosomal or alphoid sequences, as deduced from the banding and in situ hybridization results, and breaks in two seemingly unrelated regions on the two different chromosomes. The use of conventional cytogenetic techniques together with molecular studies allowed more precise evaluation of the breakpoints involved in Robertsonian translocations than either approach alone might have done.

A protein of Mr 80,000 is associated with the nucleolus organizer of human cell lines

A rabbit serum which had previously been reported to have an immunological affinity for centrosomes of human cell lines was shown also to be specific for the nucleus. Optical and ultrastructural immunolocalization in HeLa cells showed that this specificity is restricted to the fibrillar centre of nucleoli either in untreated or actinomycin D treated interphase cells. In mitotic cells discrete labelling was observed on chromosomes and shown to correspond, on spread metaphase plates, to the short arms of acrocentric chromosomes, i.e. to the nucleolar organizer regions (NORs). Using independent cell fractionation procedures in the human T-lymphoblastic KE 37 cell line and purification of immunoglobulins by affinity to antigens detected by electrophoresis and blotting, a strict correlation between immunoreactive proteins and cytological staining was established. The nucleolar specificity was shown to correspond to a protein with an Mr of 80,000 while the centrosomal specificity corresponded principally to a protein doublet of 60,000-65,000. These antigens share common epitopes as shown by the staining of both NOR and centrosome by immunoglobulins purified by affinity to either type of protein.

The technical variables associated with the frequencies of QFQ, RFA and CBG heteromorphisms of human chromosomes

In the 1971 Paris Conference (1972) it was established that certain regions of human chromosomes show remarkable heteromorphisms. These are the short arm of acrocentric chromosomes, the secondary constriction regions of chromosomes 1, 9, 16, and the distal 2/3rd of the long arm of the Y and the centromere of chromosomes 3, 4, and 5. There are several technical variables which affect the frequency of these heteromorphisms. These include quality of culture, age of slide, photography (filter, etc.), method of printing and method of scoring (criteria). Several other variables in the production of QFQ, RFA, and CBG heteromorphisms are discussed. In order to compare results from different laboratories these variables must be taken into consideration.