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Identification of three promoters in the human holocarboxylase synthetase (HCS) gene
Author(s) -
Xia Mengna,
Malkaram Sridhar Achary,
Zempleni Janos
Publication year - 2012
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.26.1_supplement.647.1
Subject(s) - promoter , enhancer , hek 293 cells , biology , gene , chromatin , genetics , human genome , repressor , genome , microbiology and biotechnology , transcription factor , gene expression
HCS catalyzes the binding of biotin to carboxylases, which play essential roles in macronutrient metabolism. HCS also appears to play a role in a multiprotein repressor complex in human chromatin. Little is known about the transcriptional regulation of HCS. This study sought to identify and characterize human HCS promoter(s). An in silico search (ensembl genome browser) revealed 3 potential promoters, positioned at bp 2888 to 5253 (P1), 12877 to 14926 (P2), and 28055 to 29336 (P3) in the HCS gene ( NG_016193 ). The promoters were cloned from human IMR90 cells and subcloned into the pGL3 luciferase reporter plasmid to create HCS‐P1GL3, HCS‐P2GL3, and HCS‐P3GL3. All 3 constructs exhibited promoter activity when transfected into human HEK293 kidney cells, HCT116 colon cells, and MCF7 mammary cells. Typically, promoter activities followed the pattern P1 > P3 > P2, but some tissue specificity was apparent. For example, the activity of P3 was about 3 times the activities of P1 and P2 in MCF7 cells. Analysis of the Encode database ( genome.ucsc.edu ) revealed a significant enrichment of the activation mark H3K4me3, suggesting that HCS is transcribed from all 3 promoters. Both enhancers and repressors were identified in all 3 promoters. We conclude that we have identified novel, functional promoters for human HCS, and that P1 is the strongest promoter in most tissues. (ARD Hatch Act, NIH DK063945 , DK077816 , and DK082476.)