THE ROLE OF ZF5 AND CGGBP-20 TRANSCRIPTION FACTORS IN EXPRESSION REGULATION OF HUMAN FMR1 GENE RESPONSIBLE
FOR X-FRAGILE SYNDROME
P. V. Gulyy,1 S. V. Orlov,1,2 E. B. Dizhe,2 K. B. Kuteikin-Teplyakov,2,3
I. A. Ignatovich,2 S. V. Zhuk,1 A. P. Perevozchikov 1,2,*
1 St. Petersburg State University, Embryology Department, 2 Institute of Experimental Medicine RAMS, St. Petersburg,
and 2 Ruhr University, Department of Neurobiochemistry, Bohum, Germany;
* e-mail: app@iem.sp.ru
The human FMR1 gene encodes an RNA-binding protein taking part in translation regulation. The 5'-untranslated region of FMR1 gene contains
a large number of tandem repeats of GCC triplets (5-50) which increasing (more then 200) is responsible for X-fragile syndrome (human congenital anomaly).
As it has been shown earlier, al least two transcription factors (ZF5 and CGGBP-20) are capable of interacting specifically with GCC-repeats in regulatory regions
of some genes. In this work, their roles in FMR1 gene expression regulation were studied. It was demonstrated by electrophoretic mobility shift assay that
ZF5 recombinant protein specifically bound with GCC-triplet repeats (GCC9). Tissue-specific distributions of ZF5 and FMR1 proteins are very overlapped
in mammalian. Inhibition of ZF5 expression in HepG2 cells (by RNA interference) leads to at least 1.5 times stimulations of FMR1 gene expression in these cells.
To estimate the contribution of GCC-triplet repeats in FMR1 gene expression regulation we used two alternative variants of genetic construction: containing
luciferase reporter gene under 5'-regulatory region fragment devoid of GCC-triplet repeats or including the GCC9 nucleotide sequence. HepG2 cells were
co-transfected by these constructions and expressions vectors of ZF5 or (and) CGGBP-20 respectively. It was found that ZF5 downregulated the activity of 5'-regulatory
region of FMR1 gene in both cases (acting probably through canonic 5'-GCGCGC3' sites). The presence of GCC-triplet repeats in the construction weakens this
ZF5 effect. CGGBP-20 downregulates the activity of 5'-region of FMR1 gene in the presence of GCC-triplets only. The data obtained evidently indicate differently
directed ZF5 effects on FMR1 gene expression and suggest the mechanism to explain the earlier demonstrated phenomenon about increasing of mRNA level in
permutation FMR1 allele carries.
Key words: GCC-triplet repeats, X-fragile syndrome, human FMR1 gene expression regulation, ZF5 and CGGBP-20 transcription factors
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