Characterization and Functional Analysis of HMG Proteins in Plasmodium falciparum

Abstract

Little is known about regulation of gene expression in Plasmodium. Indeed, only few regulatory elements and no clearly defined transcription factors have been reported, so far. In order to evaluate the interplay between cis‐ and trans‐acting elements controlling the regulation of transcription, we annotated several High Mobility Group proteins of the HMGB superfamily containing one or two HMG‐box(es) of 80 aa consisting of three α‐helices arranged in L‐shaped configuration. These computational analyses suggest that the Plasmodium proteins (PfHMG1‐4) might function as architectural factors through their ability to bind DNA in a structure‐specific manner and to induce bending of DNA. PfHMG1 (97 aa) was cloned from the genomic DNA of the 3D7 clone. The protein was expressed in E. coli and antibodies were raised in mice. A messenger of 1.3 kb was characterised by Northern blotting of 3D7 total RNA. Temporal expression of the transcript and protein was examined in two Plasmodium clones, 3D7 and F12 its gametocyte‐less derivative. The transcript appeared to be developmentally regulated throughout the erythrocytic cycle of Plasmodium as well as the protein present in the nuclear extracts. The functional analysis of the recombinant PfHMG1 via EMSA with synthetic fourway DNA junctions and ligase‐mediated circularisation assays showed that the factor is a genuine architectural protein. In addition, the functional analyses of two other factors PfHMG3 and PfHMG4 are underway. The sequences of PfHMG1 and PfHMG3 are quite similar; however, pfhmg1 transcript is mainly expressed during erythrocytic asexual cycle in contrast to pfhmg3 preferentially expressed in the gametocytes.