Clinical characterization of a large Caribbean Hispanic family linked to chromosome 9 without ApoE4

To identify the novel receptor tyrosine kinases (RTKs) critical to the proliferation of hematopoietic stem cells, we performed polymerase chain reaction-based cloning from highly purified murine hematopoietic stem cells. Lineage marker-negative, c-KIT-positive, and Ly6A/E- or Sca- 1-positive (Lin-c-KIT+Sca-1+) cells were sorted by a fluorescence- activated cell sorter. Two sets of degenerate oligonucleotide primers were directed to the conserved sequences of the catalytic domain, and were used to amplify cDNAs that encode protein tyrosine kinases (PTKs). One hundred cDNA clones were sequenced and 8 RTKs were identified, as well as 12 non-RTKs and 2 serine/threonine kinases. Sixteen cDNAs were identical to the known kinase genes (PKC beta, JAK-1, JAK-2, TYK-2, HCK, FGR, FYN, BLK, c-FES, FER, c-ABL, c-KIT, FLK-1, FLK-2, IGF1R, and ECK). Six novel cDNA sequences (stk series) were identified. However, three of them turned out to be BPK, RYK, and TEK. The remaining three showed high homology to S6 kinase II, JAK-2, and v-SEA/c-MET, respectively. Characterization of full-length cDNA sequence of the v- SEA/cMET-related gene showed that this was a novel RTK gene and we named this gene STK (stem cell-derived tyrosine kinase). We identified two distinct forms of STK cDNA; the short one encoded a putative truncated protein that lacked most of the extracellular domain. STK was expressed at various stages of hematopoietic cells, including stem cells, but we could not detect any apparent expression in other adult tissues. The expression of the truncated form of mRNA was more predominant than that of the complete form. STK was assigned by fluorescent in situ hybridization to the R-positive F1 band of chromosome 9, the same region to which hepatic growth factor-like protein has been assigned. Characterization of these PTKs, including STK, will be helpful to elucidate the molecular mechanism of the growth regulation of hematopoietic stem cells.

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Molecular cloning of a novel receptor tyrosine kinase gene, STK, derived from enriched hematopoietic stem cells

Blood ◽

10.1182/blood.v83.11.3160.3160 ◽

1994 ◽

Vol 83 (11) ◽

pp. 3160-3169 ◽

Cited By ~ 100

Author(s):

A Iwama ◽

K Okano ◽

T Sudo ◽

Y Matsuda ◽

T Suda

Keyword(s):

Stem Cells ◽

Tyrosine Kinase ◽

Hematopoietic Stem Cells ◽

Tyrosine Kinases ◽

Growth Regulation ◽

Chromosome 9 ◽

Cdna Clones ◽

Hematopoietic Stem ◽

Degenerate Oligonucleotide

Abstract To identify the novel receptor tyrosine kinases (RTKs) critical to the proliferation of hematopoietic stem cells, we performed polymerase chain reaction-based cloning from highly purified murine hematopoietic stem cells. Lineage marker-negative, c-KIT-positive, and Ly6A/E- or Sca- 1-positive (Lin-c-KIT+Sca-1+) cells were sorted by a fluorescence- activated cell sorter. Two sets of degenerate oligonucleotide primers were directed to the conserved sequences of the catalytic domain, and were used to amplify cDNAs that encode protein tyrosine kinases (PTKs). One hundred cDNA clones were sequenced and 8 RTKs were identified, as well as 12 non-RTKs and 2 serine/threonine kinases. Sixteen cDNAs were identical to the known kinase genes (PKC beta, JAK-1, JAK-2, TYK-2, HCK, FGR, FYN, BLK, c-FES, FER, c-ABL, c-KIT, FLK-1, FLK-2, IGF1R, and ECK). Six novel cDNA sequences (stk series) were identified. However, three of them turned out to be BPK, RYK, and TEK. The remaining three showed high homology to S6 kinase II, JAK-2, and v-SEA/c-MET, respectively. Characterization of full-length cDNA sequence of the v- SEA/cMET-related gene showed that this was a novel RTK gene and we named this gene STK (stem cell-derived tyrosine kinase). We identified two distinct forms of STK cDNA; the short one encoded a putative truncated protein that lacked most of the extracellular domain. STK was expressed at various stages of hematopoietic cells, including stem cells, but we could not detect any apparent expression in other adult tissues. The expression of the truncated form of mRNA was more predominant than that of the complete form. STK was assigned by fluorescent in situ hybridization to the R-positive F1 band of chromosome 9, the same region to which hepatic growth factor-like protein has been assigned. Characterization of these PTKs, including STK, will be helpful to elucidate the molecular mechanism of the growth regulation of hematopoietic stem cells.

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Characterization of the porcine uncoupling proteins 2 and 3 (UCP2 & UCP3) and their localization to chromosome 9 p by somatic cell hybrids

Animal Genetics ◽

10.1046/j.1365-2052.1999.00462.x ◽

1999 ◽

Vol 30 (3) ◽

pp. 221-224 ◽

Cited By ~ 15

Author(s):

P. Werner ◽

S. Neuenschwander ◽

G. Stranzinger

Keyword(s):

Somatic Cell ◽

Uncoupling Proteins ◽

Chromosome 9 ◽

Somatic Cell Hybrids ◽

Cell Hybrids

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Generation and characterization of a human chromosome 9 cosmid library

Somatic Cell and Molecular Genetics ◽

10.1007/bf01233863 ◽

1992 ◽

Vol 18 (3) ◽

pp. 269-284 ◽

Cited By ~ 3

Author(s):

Sharon L. Graw ◽

Alan J. Buckler ◽

Deborah E. Britt ◽

Cynthia L. Jackson ◽

Domenica Taruscio ◽

...

Keyword(s):

Human Chromosome ◽

Chromosome 9 ◽

Cosmid Library

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Cloning and characterization of mouse extracellular-signal-regulated protein kinase 3 as a unique gene product of 100 kDa

Biochemical Journal ◽

10.1042/bj3460169 ◽

2000 ◽

Vol 346 (1) ◽

pp. 169-175 ◽

Cited By ~ 20

Author(s):

Benjamin TURGEON ◽

Marc K. SABA-EL-LEIL ◽

Sylvain MELOCHE

Keyword(s):

Protein Kinase ◽

Gene Product ◽

Mammalian Cells ◽

Chromosome 9 ◽

Mouse Tissue ◽

Mouse Development ◽

Extracellular Signal ◽

Mouse Tissues

MAP (mitogen-activated protein) kinases are a family of serine/threonine kinases that have a pivotal role in signal transduction. Here we report the cloning and characterization of a mouse homologue of extracellular-signal-regulated protein kinase (ERK)3. The mouse Erk3 cDNA encodes a predicted protein of 720 residues, which displays 94% identity with human ERK3. Transcription and translation of this cDNA in vitro generates a 100 kDa protein similar to the human gene product ERK3. Immunoblot analysis with an antibody raised against a unique sequence of ERK3 also recognizes a 100 kDa protein in mouse tissues. A single transcript of Erk3 was detected in every adult mouse tissue examined, with the highest expression being found in the brain. Interestingly, expression of Erk3 mRNA is acutely regulated during mouse development, with a peak of expression observed at embryonic day 11. The mouse Erk3 gene was mapped to a single locus on central mouse chromosome 9, adjacent to the dilute mutation locus and in a region syntenic to human chromosome 15q21. Finally, we provide several lines of evidence to support the existence of a unique Erk3 gene product of 100 kDa in mammalian cells.

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Differentiating between Hyperdiploidy and Pseudo-Hyperdiploidy in B-Lymphoblastic Leukemia Utilizing Low-Coverage Mate-Pair Sequencing

Blood ◽

10.1182/blood-2019-122440 ◽

2019 ◽

Vol 134 (Supplement_1) ◽

pp. 5212-5212

Author(s):

James B Smadbeck ◽

Beth A Pitel ◽

Kathryn E. Pearce ◽

Sarah H Johnson ◽

Nicole Hoppman ◽

...

Keyword(s):

Genetic Characterization ◽

Conflicts Of Interest ◽

Lymphoblastic Leukemia ◽

Neutral Loss ◽

Chromosome 9 ◽

Chromosomal Microarray ◽

Mate Pair ◽

Low Coverage ◽

Base Coverage

Purpose: B-cell acute lymphoblastic leukemia/lymphoma (B-ALL/LBL) represents the most common childhood malignancy. Classification of recurrent genetic abnormalities in B-ALL/LBL is essential for risk stratification and patient management. Observed in approximately 5% of B-ALL/LBL cases, hypodiploidy is considered a poor prognostic finding and can be further characterized as near-haploidy (24-31 chromosomes), low hypodiploidy (32-39 chromosomes) and high hypodiploidy (40-43 chromosomes). Importantly, masked hypodiploid clones that have undergone endoreduplication can be mischaracterized as hyperdiploidy, a favorable prognostic finding in B-ALL/LBL. These "doubled" hypodiploid clones are termed pseudo-hyperdiploid and their accurate detection is critical. Mate-pair sequencing (MPseq) is a next-generation sequencing (NGS) technology designed to sequence larger DNA fragments (2000-5000bp) than paired-end sequencing. This allows for the detection of large genetic rearrangements (e.g. translocations, inversions, etc.) and copy number variants (CNVs) at a high resolution with low base coverage. It has demonstrated particular clinical utility for genetic characterization of hematologic malignancies. To expand the role of MPseq in the evaluation of hematologic neoplasms, we aimed to improve the analysis of MPseq results to allow for the detection of copy-neutral loss of heterozygosity (cnLOH) that would enable the differentiation between hyperdiploid and pseudo-hyperdiploid clones. Methods: A new algorithm was applied that statistically analyzes common SNP locations in MPseq data in order to detect both cnLOH regions, and provides global heterozygosity levels at a resolution that allows for the detection of hypodiploidy. This method accounts for the low base-coverage of MPseq data (<10x) in order to provide the highest possible accuracy and precision without increasing cost by requiring changes to the input or sequencing requirements. The algorithm was applied to 17 B-ALL samples classified by conventional chromosome analysis, FISH, and/or chromosomal microarray as either pseudo-hyperdiploidy (n=8) or hyperdiploidy (n=9). The cases were sequenced using MPseq with 101bp read lengths across a range of base coverages (1-10x; 30-135 million fragments) and tumor percentages (50-100%). Results: We demonstrate the method's ability to differentiate between hyperdiploidy and pseudo-hyperdiploidy in all cases without any changes to the MPseq input or sequencing requirements. We detected an average of 11.5 whole-chromosome cnLOH (range 8-20) in the pseudo-hyperdiploidy cases, compared to an average of 0.67 whole-chromosome cnLOH (range 0-2) in the hyperdiploidy cases. Additionally, the method allows for the detection of cnLOH regions across the genome. For example, we observed a recurrent cnLOH of chromosome 9 in the hyperdiploid subtype (3/9 hyperdiploid cases), which is a recurrent abnormality in the disease. The method provides an easy to read visual output that compares favorably to the output of chromosomal microarray, increasing its utility for clinical application. Conclusions: This advancement in the analysis of low-coverage sequencing data makes MPseq available for the genetic characterization of B-ALL/LBL, where hypodiploidy is a primary cytogenetic abnormality and is sometimes missed by conventional cytogenetic techniques. Additionally, the algorithm allows for the detection of cnLOH regions across the genome, which is important across many disease types for diagnosis, prognosis, and the detection of potential therapeutic targets. The addition of the algorithm to the MPseq analysis pipeline makes it a more complete genetic characterization tool and brings it closer to its promise as a potential single assay replacement for conventional cytogenetic techniques. Disclosures No relevant conflicts of interest to declare.

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