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S N Sait

Publications and source records attributed to S N Sait.

At least 19 recordsLinked to original sources
PubMed2005

Alternative mechanisms associated with silencing of CDKN1C in Beckwith-Wiedemann syndrome.

BACKGROUND: Mutations in the imprinted gene CDKN1C account for approximately 10% of Beckwith-Wiedemann syndrome (BWS) cases. Fibroblasts from BWS patients with loss of methylation (LOM) at the imprinting control region (ICR) KvDMR1 have reduced CDKN1C expression. Another group of BWS patients with downregulated CDKN1C expression but with normal methylation at KvDMR1 has been identified. OBJECTIVE: To investigate the mechanism of CDKN1C silencing in BWS in these two classes of patients. METHODS: The CDKN1C promoter region was analysed for changes in DNA methylation using bisulphite sequencing, and for alterations in chromatin structure using the chromatin immunoprecipitation (ChIP) assay. RESULTS: There was only spurious CpG methylation of the CDKN1C promoter in fibroblast DNA from both normal individuals and patients with BWS, irrespective of the methylation status of KvDMR1. There was no detectable change in chromatin structure at the CDKN1C promoter in patients with LOM at KvDMR1. BWS patients with downregulated CDKN1C and normal methylation at KvDMR1 had depletion of dimethylated H3-K4 and enrichment of dimethylated H3-K9 and HP1gamma at the CDKN1C promoter, suggesting that in these cases gene silencing is associated with repressive chromatin changes. CONCLUSIONS: CDKN1C may be downregulated by multiple mechanisms including some that do not involve promoter methylation. In BWS patients with normal methylation at KvDMR1 and reduced expression of CDKN1C, repressive chromatin may play a role, but the absence of methylation and repressive chromatin structure at the CDKN1C promoter in BWS patients with LOM at KvDMR1 argues for a direct role of this epimutation in silencing CDKN1C.

Beckwith-Wiedemann Syndrome↗
PubMed2001

Trisomy 6 in basal cell carcinomas correlates with metastatic potential: a dual color fluorescence in situ hybridization study on paraffin sections.

BACKGROUND: Most basal cell carcinomas (BCCs) are indolent lesions; a few become locally aggressive or even metastatic. Little is known about the molecular and genetic alterations in this malignant transformation. Conventional karyotyping in BCC has revealed a high frequency of nonclonal, structural rearrangements, with few cases that show multiple, unrelated, small clones suggestive of a multicellular origin. Trisomy 6 was described recently in a few BCCs, but the biologic significance of the appearance of trisomy 6 in BBCs was not clear. METHODS: Thirty cases including 4 metastatic, 4 locally aggressive, and 22 conventional nonaggressive BCCs were studied. Fluorescence in situ hybridization (FISH) was performed on 4 microm tissue sections, using alpha-centromeric enumeration probes for chromosome 6 (SpectrumGreen, Vysis Inc., Downers Grove, IL) and chromosome 4 (SpectrumOrange, Vysis Inc., Downers Grove, IL, used as disomic cell control). Trisomy 6 was semiquantitated within tumor cells and nonneoplastic cells in each case. RESULTS: Trisomy 6 was identified in all 4 metastatic BCCs within tumor cells and in corresponding BCCs at the primary cutaneous site in 2 of these 4 cases. Two locally aggressive BCCs, 1 of which had preceding radiation exposure, also showed trisomy 6. All nonaggressive BCCs and nonneoplastic cells were disomic for chromosome 6. CONCLUSIONS: Trisomy 6 has been identified as a cytogenetic aberration representative of tumor cells in aggressive and metastatic BCC. None of the nonaggressive BCCs in this study demonstrated trisomy 6. Acquisition of trisomy 6 by tumor cells in BCC may lead to the emergence of metastatic potential. Additional studies to define the underlying mechanisms may be valuable in preventing aggressive behavior in BCC.

Adult↗
PubMed2001

High-dose cytosine arabinoside and idarubicin treatment of chronic myeloid leukemia in myeloid blast crisis.

Chronic myeloid leukemia in myeloid blast crisis (CML-MBC) is highly resistant to standard induction chemotherapy regimens. Anecdotal results from previous clinical trials support the concept of dose escalation in patients with CML-MBC. Eight patients with CML-MBC were treated with cytosine arabinoside (Ara-C) 1.5-3.0 g/m2 intravenously over 1 hr every 12 hr for 12 doses and idarubicin 12 mg/m2 intravenously daily for 3 days. Sixteen previous reports describing the use of Ara-C-based chemotherapy regimens in patients with CML-MBC were also reviewed. Our patients' median age was 62 years (range, 42-69 years). One patient achieved complete hematologic remission (95% confidence interval, 0.3%, 53%). The median survival for our patients was 7.3 months. These results were not different from previous published reports using Ara-C-based chemotherapy regimens to treat CML-MBC. In summary, the combination of high-dose Ara-C and idarubicin did not improve the overall prognosis of patients with CML-MBC. Innovative approaches need to be explored for this patient population.

Adult↗
PubMed2001

Truncated STAT proteins are prevalent at relapse of acute myeloid leukemia.

Signal transducer and activator of transcription (STAT) proteins are implicated in the control of cell survival, proliferation and differentiation in response to hematopoietic cytokines. C-terminally truncated STAT isoforms (STATbeta), as opposed to the full length form (STATalpha), have a competitive or even transdominant negative effect on gene induction mediated by the STAT pathway. We have previously demonstrated that while constitutively active STAT proteins were detected in ten of 36 (28%) for STAT3 and eight of 36 (22%) for STAT5 in pretreatment samples from newly diagnosed acute myeloid leukemia (AML) patients, a significantly larger fraction of samples [21 of 27 (78%)] expressed STATbeta proteins. To determine whether STATbeta expression was maintained or increased after relapse in AML, we compared STAT activity and isoform expression at diagnosis and at relapse in 17 patients. In this selected group, constitutively active STAT3 was detected in 13 of 17 (76%) AML samples at diagnosis but was detected in only four of these patients at relapse. Constitutively active STAT5 was detected in three of 17 (18%) AML samples at diagnosis; but only two at relapse. In contrast, STATbeta protein expression was observed in 12 of the 17 pretreatment samples (71%) and in 16 of 17 samples at relapse. Only one patient did not express STATbeta at relapse. Our results suggest that STATbeta isoform expression, rather than level of constitutive activity, may be involved in disease progression in AML.

Adult↗
PubMed2001

The pericentromeric region of human chromosome 11: evidence for a chromosome-specific duplication.

We have identified a chromosome duplication in the pericentromeric region of human chromosome 11 located in 11p11 and 11q14. A detailed physical map of each duplicated region was generated to describe the nature of the duplication, the involvement at the centromere and to resolve the correct maps. All clones were evaluated to ensure they were representative of their genetic origin. The order of clones, based on their marker content, as well as the distance covered was determined by SEGMAP. Each duplication encompasses more than 1 Mb of DNA and appears to be chromosome 11 specific. Ten STS markers were mapped within each duplication. Comparative sequence analysis along the duplication identified 35 nucleotide changes in 2,036 bp between the two copies, suggesting the duplication occurred over 14 million years ago. A suggested organization of the pericentromeric region, including the duplications and alpha-related repetitive sequences, is presented.

Centromere↗
PubMed2001

Acute myeloid leukemia in the setting of low dose weekly methotrexate therapy for rheumatoid arthritis.

Methotrexate is in widespread use as second-line therapy for rheumatoid arthritis. Treatment with methotrexate in this and other settings has not been associated with the development of therapy-related leukemias. Four patients with rheumatoid arthritis are reported who developed acute myeloid leukemia (AML) while receiving low dose weekly methotrexate therapy in the absence of previous or concomitant treatment with known leukemogenic agents. AML in these four patients was of different morphologic subtypes and was associated with heterogeneous cytogenetic abnormalities, cell surface marker expression and multidrug resistance protein expression. None of the recognized features of therapy-related leukemia were present in these four nor in five previously-reported patients. It is likely that the occurrence of AML in patients with rheumatoid arthritis in the setting of methotrexate therapy represents the coincidence of these two diseases, and does not reflect a causal relationship.

Aged↗
PubMed1999

Demonstration of urokinase expression in cancer cells of colon adenocarcinomas by immunohistochemistry and in situ hybridization.

The question whether urokinase is expressed in human colon cancer by the cancer cells themselves or by surrounding stromal elements such as fibroblasts, macrophages, and leukocytes, which transfer the activator to the receptors of the cancer cells, has been a controversial one. In the present study 12 cases of colorectal cancer were investigated by immunohistochemical methods using three monoclonal antibodies of different specificity against urokinase. Cytoplasmic staining of strongly varying intensity was observed in all cases, with the antigen expressed most strongly in the apical and the basal regions of the cancer cells. In some cases, staining was also found in stromal elements surrounding the cancer glands. That the activator was indeed the product of the cancer cells was demonstrated by in situ hybridization using a uPA-cDNA probe, which detected the presence of uPA-mRNA in both the basal and the apical regions of the cancer cells. A monoclonal antibody against the receptor for uPA showed similar localization. These findings indicate that the activator is expressed by the cancer cells and is not recruited by them from stromal elements.

Adenocarcinoma↗
PubMed1997

A 1-Mb physical map and PAC contig of the imprinted domain in 11p15.5 that contains TAPA1 and the BWSCR1/WT2 region.

We have constructed a 1-Mb contig in human chromosomal band 11p15.5, a region implicated in the etiology of several embryonal tumors, including Wilms tumor, and in Beckwith-Wiedemann syndrome. Cosmid, P1, PAC, and BAC clones were characterized by NotI/SalI digestion and hybridized to a variety of probes to generate a detailed physical map that extends from D11S517 to L23MRP. Included in the map are the CARS, NAP2, p57/KIP2, KVLQT1, ASCL2, TH, INS, IGF2, H19, and L23MRP genes as well as end probes isolated from PACs. The TAPA1 gene, whose protein product can transmit an antiproliferative signal, was also localized in the contig. However, Northern blot analysis demonstrated that its expression did not correlate with tumorigenicity in G401 Wilms tumor hybrids, suggesting that TAPA1 is not responsible for the tumor suppression associated with 11p15.5. Genomic clones were used as probes in FISH analysis to map the breakpoints from three Beckwith-Wiedemann syndrome patients and a rhabdoid tumor. Interestingly, each of the breakpoints disrupts the KVLQT1 gene, which is spread over a 400-kb region of the contig. Since 11p15.5 contains several genes with imprinted expression and one or more tumor suppressor genes, our contig and map provide a framework for characterizing this intriguing genetic environment.

Antigens, CD↗
PubMed1997

The human HNP36 gene is localized to chromosome 11q13 and produces alternative transcripts that are not mutated in multiple endocrine neoplasia, type 1 (MEN I) syndrome.

Multiple endocrine neoplasia, type 1 (MEN I), is an autosomal dominant syndrome of selected endocrine neoplasms whose causative gene, a suspected tumor suppressor, has been localized to chromosome 11q13, but has not been identified. Recently, the HNP36 cDNA was identified as a novel growth factor responsive gene of undetermined biological function that is expressed in the pituitary and parathyroid glands. In studies seeking the function of the HNP36 gene product, the gene was localized by fluorescence in situ hybridization within the 11q13 segment. Further analysis of radiation-reduced hybrid DNAs and chromosome 11-specific YAC clones established that the HNP36 gene is within 80 kb of D11S913, a marker tightly linked to the MEN1 gene. Consequently, the HNP36 gene was studied as a candidate for the MEN1 gene. The human HNP36 gene was cloned and determined to consist of 12 exons. Expression of the HNP36 gene from pituitary and parathyroid tissue and four patient tumors or lymphoblasts was confirmed by RT-PCR amplification of the coding sequences, and HNP36 transcripts were analyzed for mutations. All tissues expressed three HNP36 gene transcripts that result from alternative splicing and appear to encode related, but distinct, proteins. However, DNA sequence determination of the RT-PCR products from MEN I-associated tumors found no deletions and identified a single nucleotide difference that may be a polymorphism. Thus, mutations in the coding segments of the HNP36 gene are not the cause of the MEN I syndrome. Nevertheless, the assignment of the HNP36 gene to 11q13 and identification of new potential gene products provides a novel growth-regulated genetic candidate for other disorders whose genes map to this locus.

Alternative Splicing↗
PubMed1997

Chromosomal localization of a human mucin gene (MUC8) and cloning of the cDNA corresponding to the carboxy terminus.

A partial cDNA (pAM1) encoding a major airway mucin glycoprotein with novel tandem repetitive sequence has recently been cloned (Shankar, V., M. S. Gilmore, R. C. Elkins, and G. P. Sachdev. 1994. Biochem. J. 300:295-298). In this article, we report additional new sequence derived by 3'-rapid amplification of cDNA ends technique. The sequence corresponds to a stop codon, 3'-untranslated region of 458 bp, a polyadenylation signal, and poly A+ tail, and represents the extreme carboxy terminus of MUC8. A plasmid construct (pAM3) in pBluescript was generated by in-frame ligation of pAM1 to the 479-bp 3'UTR of MUC8. A 5'-end 325-bp fragment of this cDNA subcloned into the protein fusion and expression vector pET28b(+) was used to generate fusion protein under the control of T7 promoter. The purified fusion protein as well as synthetic peptide corresponding to the MUC8 repeat sequence (TSCPRPLQEGTPGS) were used to raise polyclonal antibodies in rabbits. The antiserum to the fusion protein and to the synthetic peptide reacted with the deglycosylated major tracheobronchial mucin. Immunohistochemical studies using the above antibodies localized the MUC8 protein product to submucosal glands in human tracheal epithelium. Furthermore, the gene from which this cDNA is derived, was mapped to chromosome 12 using DNA from a panel of human-mouse somatic cell hybrids. Fluorescence in situ hybridization was used to assign the regional localization to 12q24.3. Since the eight known human mucin genes map to other chromosomes, we have named this gene MUC8, in accordance with mucin gene nomenclature.

Amino Acid Sequence↗
PubMed1996

A high-resolution physical map of human chromosome 11.

The development of a highly reliable physical map with landmark sites spaced an average of 100 kbp apart has been a central goal of the Human Genome Project. We have approached the physical mapping of human chromosome 11 with this goal as a primary target. We have focused on strategies that would utilize yeast artificial chromosome (YAC) technology, thus permitting long-range coverage of hundreds of kilobases of genomic DNA, yet we sought to minimize the ambiguities inherent in the use of this technology, particularly the occurrence of chimeric genomic DNA clones. This was achieved through the development of a chromosome 11-specific YAC library from a human somatic cell hybrid line that has retained chromosome 11 as its sole human component. To maximize the efficiency of YAC contig assembly and extension, we have employed an Alu-PCR-based hybridization screening system. This system eliminates many of the more costly and time-consuming steps associated with sequence tagged site content mapping such as sequencing, primer production, and hierarchical screening, resulting in greater efficiency with increased throughput and reduced cost. Using these approaches, we have achieved YAC coverage for >90% of human chromosome 11, with an average intermarker distance of <100 kbp. Cytogenetic localization has been determined for each contig by fluorescent in situ hybridization and/or sequence tagged site content. The YAC contigs that we have generated should provide a robust framework to move forward to sequence-ready templates for the sequencing efforts of the Human Genome Project as well as more focused positional cloning on chromosome 11.

Chimera↗
PubMed1996

Structure and chromosomal localization of the human salivary mucin gene, MUC7.

We have isolated and characterized several MUC7 genomic clones encoding the human low-molecular-weight salivary mucin, MG2. The MUC7 gene spans approximately 10.0 kb and comprises of three exons and two introns. Intron 1 is approximately 1.7 kb long and is located in the 5'-untranslated region of the corresponding MUC7 cDNA. Intron 2 spans approximately 6.0 kb and is located close to the boundary of the putative leader peptide and secreted protein. The entire region encoding the secreted peptide is located on exon 3, spanning approximately 2.2 kb. The nucleotide sequence of sections of the MUC7 gene, including 1500 bp of the 5'-flanking region, was determined and analyzed for motifs identical or homologous to other known response elements. A modified RACE procedure was used to determine the 5'-end of the MUC7 mRNA. PCR, the human-hamster somatic cell hybrid panel PCRable DNAs kit, and an in situ hybridization analysis on the complete metaphase chromosome spreads were used for the chromosomal localization of the MUC7 gene. It was mapped to chromosome 4q13-q21.

Animals↗
PubMed1996

ML-1 cell line lacks a germline MLL locus.

Gene rearrangements involving MLL (also known as ALL1, HRX, or Htrx) are among the most common molecular abnormalities associated with acute leukemia. These leukemias generally have one allele involved in a rearrangement, while the remaining allele is uninvolved and demonstrates a germline MLL configuration. In this study, we describe a leukemic cell line that does not have a germline MLL allele and thus cannot produce a normal MLL gene product. We show that the ML-1 cell line, derived from a patient with acute myeloid leukemia, has one allele involved in a t(6;11)(q27;q23), while the remaining MLL allele is deleted. Cloning of the genomic breakpoints on the derivative(6) and der(11) chromosomes demonstrated a balanced translocation between MLL on chromosome band 11q23 and AF6 on chromosome band 6q27. Sequence analysis of the derivative chromosomes revealed that a 186-bp segment of MLL intron 6, downstream of the breakpoint, had been duplicated, inverted, and inserted between MLL and AF6 on the der(11) chromosome. In light of the fact that ML-1 cells can be induced to differentiate along the granulocyte and macrophage lineages, the finding that ML-1 lacks a germline MLL allele demonstrates that a normal MLL gene is not required for survival, proliferation, or differentiation of this cell line.

Adult↗
PubMed1995

Genomic organization and chromosomal localization of the gene encoding human P-selectin glycoprotein ligand.

The gene for P-selectin glycoprotein ligand (PSGL-1) has been cloned from a human placenta genomic DNA library. A single intron of approximately 9 kilobases was found in the 5'-untranslated region and the complete coding region resides in exon 2. The genomic clone differs from the cDNA clone isolated from HL-60 cells in that it encodes an extra copy of the decameric repeat located in the extracellular domain of PSGL-1. Further analysis indicated that the PSGL-1 genes of HL-60 and U-937 cells contain 15 repeats, whereas the PSGL-1 genes of polymorphonuclear leukocytes, monocytes, and several other cell lines contain 16 repeats. Transfection experiments did not indicate a functional difference between these two variants of PSGL-1. The two previously observed PSGL-1 mRNA species of 2.5 and 4 kilobases most likely arise from differential utilization of polyadenylation signal sequences. The organization of the PSGL-1 gene closely resembles those of CD43 and human platelet glycoprotein GPIb alpha, both of which have an intron in the 5'-noncoding region, a long second exon containing the complete coding region, and TATA-less promoters. The gene for human PSGL-1, which has been designated SELPLG by the Human Gene Nomenclature Committee, was mapped to chromosome 12q24 using Southern blot analysis of DNA from a set of human-mouse cell hybrids, and fluorescent in situ hybridization on metaphase chromosome spreads.

Amino Acid Sequence↗
PubMed1995

Chromosomal localization of the gene encoding the human DNA helicase RECQL and its mouse homologue.

We have determined the chromosomal location of the human and mouse genes encoding the RECQL protein, a putative DNA helicase homologous to the bacterial DNA helicase, RecQ. RECQL was localized to human chromosome 12 by analysis of human-rodent somatic cell hybrid DNA; fine mapping of RECQL by fluorescence in situ hybridization revealed its chromosomal location to be 12p11-p12. The corresponding mouse gene, Recql, was mapped to the telomeric end of mouse chromosome 6 by analysis of DNA from an interspecific cross.

Adenosine Triphosphatases↗
PubMed1995

Mapping of ribosomal protein S3 and internally nested snoRNA U15A gene to human chromosome 11q13.3-q13.5.

The mammalian ribosome is a massive structure composed of 4 RNA species and about 80 different proteins. One of these ribosomal proteins, S3, appears to function not only in translation but also as an endonuclease in repair of UV-induced DNA damage. Moreover, the first intron of human RPS3 transcripts is processed to generate U15A, a small nucleolar RNA. We localized the nested RPS3/U15A genes to the immediate vicinity of D11S356 and D11S533 on human chromosome 11q13.3-q13.5 using a combination of somatic cell hybrid analysis, fluorescence in situ hybridization, and YAC/STS content mapping. These findings add to the evidence that genes encoding ribosomal proteins are scattered about the human genome.

Animals↗
PubMed1995

Complex MLL rearrangement in a patient with T-cell acute lymphoblastic leukemia.

MLL (also known as ALL-I, HTRX, or HRX) gene translocations are among the most common chromosomal abnormalities recognized in both B-lineage acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML). However, MLL gene rearrangements are uncommon in T-cell ALL. We recently detected an MLL gene rearrangement in a patient with typical T-cell ALL. We recently detected an MLL gene rearrangement in a patient with typical T-cell ALL (CD2+, CD4+, CD5+, CD7+, CD8+, HLA DR-) and an apparently normal karyotype (46,XX). The rearrangement was cloned and characterized; a DNA fragment distal to the breakpoint was mapped by fluorescence in situ hybridization (FISH) to 19p13, indicating that the leukemic blasts had undergone a cytogenetically undetected rearrangement involving chromosomes 11 and 19. A reverse transcriptase-polymerase chain reaction (RT-PCR) assay demonstrated an in-frame fusion mRNA between the amino terminus of MLL and the carboxy terminus of ENL (also known as MLLT1 or LTG19), a gene that has been mapped to 19p13. In addition, MLL sequences distal (telomeric) to the breakpoint were deleted from the genome, which precludes the formation of a reciprocal ENL/MLL fusion protein. These findings suggest that an MLL/ENL fusion protein (and not a reciprocal ENL/MLL fusion) was likely to be pathogenic in this patient, and they reinforce previous studies showing that leukemic blasts with apparently normal karyotype may harbor MLL rearrangements. Additionally, this report provides the first conclusive evidence of an MLL/ENL gene fusion characterized at a molecular level in a patient with T-cell ALL.

Base Sequence↗