Regulation
of MHC class I genes expression level is critical to achieve proper immune
surveillance. In this work, we identify DNA sequences downstream of the
promoter, necessary for MHC class I regulation. We first demonstrate that in
transgenic mice, the promoter of MHC class I gene, in the absence of downstream
elements, is insufficient to direct expression of reporter genes. However, in
the presence of the coding and 3' flanking sequences high levels of expression
and proper tissue distribution are observed, pointing to the requirement for
elements downstream of the promoter for MHC class I regulation. We identify two
sets of elements downstream of the promoter that are necessary for appropriate
regulation: Proper tissue-specific regulation requires the presence of the
first two introns, possibly pointing to direct binding of tissue specific
factors to sequence element(s) within the proximal introns. In addition, sequences
beyond the polyA addition site are necessary for expression in vivo, but
have no effect in transient transfection assays. Using transgenic mice and
stably transfected cell lines, we demonstrate that a 3’ segment corresponding
to the untranslated and 723bp intergenic regions functions as a barrier
element, protecting the MHC class I gene from transcriptional silencing by
propagation of repressive epigenetic modifications. Accordingly, truncation of
the 3’ segment is correlated with gene silencing accompanied by repressive
chromatin organization: increased nucleosomes density and decreased histone
H3K9 acetylation and H3K4 methylation across the gene. Moreover, using
chromatin immunoprecipitation, we show association of histone modifying enzymes
with the 3' segment. Histone variant H2A.Z, which was associated both with
barrier elements and activated promoters, is also associated with the MHC class
I 3' segment, as well as the promoter. Taken together, these findings demonstrate
the existence of a novel barrier element downstream of the MHC class I polyA
addition site which functions to maintain transcription-permissive chromatin
organization.
