The human serglycin gene. Nucleotide sequence and methylation pattern in human promyelocytic leukemia HL-60 cells and T-lymphoblast Molt-4 cells.
The complete nucleotide sequence of the 16.7-kb human gene that encodes the peptide core (serglycin) of a secretory granule proteoglycan was determined, thus representing the first proteoglycan peptide core gene to be sequenced in its entirety. The exons, intron 1, and intron 2 comprised 7, 53, and 40% of the gene, respectively. Nineteen Alu-repetitive DNA sequences were interspersed in the gene, accounting for 28% of the total nucleotides in intron 1 and 40% of the nucleotides in intron 2. The nucleotide sequence was then used in an examination of the methylation pattern of the human serglycin gene in human promyelocytic leukemia HL-60 cells that contain serglycin mRNA and in T-lymphoblast Molt-4 cells that do not. With polymerase chain reaction methodology, 13 DNA probes of 250-880 base pairs in length were generated that corresponded to unique, non-Alu sequences spaced throughout the entire human serglycin gene. When blots containing genomic DNA digested with HpaII or MspI were examined with these genomic probes, it was discovered that the 5'-flanking region and intron 1 of the serglycin gene in HL-60 cells were both substantially less methylated than intron 2. In contrast, the entire serglycin gene in Molt-4 cells was highly methylated. Because hypomethylated genes generally are transcribed more efficiently than hypermethylated genes, the high level of serglycin mRNA in HL-60 cells probably is a consequence of the low level of methylation of intron 1 and the 5'-flanking region of the serglycin gene in these cells.
Humphries
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Department of Veterans Affairs Outpatient Clinic, Boston, Massachusetts.
Nicodemus
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Schiller
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Stevens
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D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D004247:DNA; D006801:Humans; D007438:Introns; D015473:Leukemia, Promyelocytic, Acute / Q000378:metabolism*; D008745:Methylation; D008969:Molecular Sequence Data; D011509:Proteoglycans / Q000235:genetics*; D012313:RNA / Q000235:genetics; D013601:T-Lymphocytes / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D033921:Vesicular Transport Proteins
D011509:Proteoglycans; D033921:Vesicular Transport Proteins; C075430:serglycin; D012313:RNA; D004247:DNA