title: Transcriptional control of the B3GALT5 gene by a retroviral promoter and methylation of distant regulatory elements.
abstract: We focused on transcription factors and epigenetic marks that regulate the B3GALT5 gene through its retroviral long terminal repeat (LTR) promoter. We compared the expression levels of the B3GALT5 LTR transcript, quantitated by competitive RT-PCR, with those of the candidate transcription factors HNF1α/β and Cdx1/2, determined by Western blot analysis, in colon cancer biopsies, various cell lines, and cell models serving as controls. We found that HNF1α/β were easily detected, irrespective of the amount of LTR transcript expressed by the source, whereas Cdx1/2 were undetectable, and no sample lacking HNF1α/β expressed the LTR transcript. On transfection in proper host cells, both HNF1α and HNF1β provided detectable LTR transcript, whereas shRNA-mediated silencing of HNF1β impaired transcription. Treating cells with 5'-aza-2'-deoxycytidine (5AZA) strongly reduced expression, without affecting HNF1α/β, despite the lack of CpG islands in the LTR and proximal sequences. By electrophoresis mobility shift and luciferase reporter assays, the LTR promoter binding and activity did not correlate with the amounts of LTR transcript expressed in the cells and depended on the levels of the transcription factors. We conclude that HNF1α/β are necessary but insufficient to activate and regulate B3GALT5 LTR transcription, which depends on unknown regulatory elements that are active when methylated and located outside of and far from the LTR promoter.
authors.affiliation: 1University of Insubria, via J. H. Dunant 5, 21100, Varese, Italy. marco.trinchera@uninsubria.it.
authors.forename: Aida
authors.initials: A
authors.lastname: Zulueta
authors.forename: Anna
authors.initials: A
authors.lastname: Caretti
authors.forename: Paola
authors.initials: P
authors.lastname: Signorelli
authors.forename: Fabio
authors.initials: F
authors.lastname: Dall'olio
authors.forename: Marco
authors.initials: M
authors.lastname: Trinchera
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D005700:Galactosyltransferases / Q000235:genetics / Q000378:metabolism*; D051538:Hepatocyte Nuclear Factor 1-alpha / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012190:Retroviridae / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D051538:Hepatocyte Nuclear Factor 1-alpha; C479886:B3GALT5 protein, human; D005700:Galactosyltransferases
keywords: colon cancer; hepatocyte nuclear factor 1; transposon; type 1 chain carbohydrates