cis-Acting Regulatory Elements in the Bovine Immunodeficiency Virus Long Terminal Repeat

Abstract

Functional cis-acting regulatory elements in the bovine immunodeficiency virus (BIV) long terminal repeat (LTR) were identified by deletion mapping and nuclear protein gel shift analysis using three BIV-infectible cell lines, Cf2Th, BLAC-20, and EREp. Deletion mapping studies indicated that putative NF-kappa B, GRE, AP-4, AP-1, CAAT, and ATF/CRE transcription factor elements positively contribute to LTR-directed gene expression in each cell line both in the presence and absence of the viral transactivator Tat. Sp1 and overlapping AP-3 and retroviral core enhancer elements had variable effects on LTR-directed gene expression depending on cell type and presence or absence of Tat. In addition, a sequence spanning the LTR U5 region and the untranslated viral leader was strongly repressive in all cell lines. Tat transactivated the LTR 25-fold over basal levels in a TAR-dependent manner in Cf2Th cells. In contrast, Tat transactivated the LTR only 2.5-fold over basal levels in EREp and BLAC-20 cells in a TAR-independent manner. Probes for putative NF-kappa B, GRE, Sp1, AP-4, AP-1, overlapping AP-3 and retroviral core enhancer, and juxtaposed CAAT and ATF-CRE elements specifically bound nuclear proteins from these three cell lines and HeLa cells, with the stoichiometry of binding being cell-type dependent. Probes for AP-4, AP-1, and juxtaposed CAAT and ATF/CRE elements exhibited greater protein binding with extracts from virally infected cells than with extracts from uninfected cells, suggesting that viral infection can modulate nuclear factor binding. The present studies indicate that several transcription factor elements in the BIV LTR have functional roles and that cell type can strongly determine the role they play in gene expression.

Binding of Tat to TAR and Recruitment of Positive Transcription Elongation Factor b Occur Independently in Bovine Immunodeficiency Virus

“…Thus, bTat brings P-TEFb to bTAR for the transcriptional needs of BIV. Moreover, we resolved the dilemma of why some cells that are permissive for BIV infection do not support bTat transactivation (13). In these cells, the bLTR is a weak promoter and probably does not synthesize sufficient amounts of bTAR transcripts.…”

“…Having established that cyclins T1 from different species bind to bTat in vitro and knowing that they are conserved from Drosophila melanogaster to Homo sapiens, we were intrigued by low levels of bTat transactivation in virally permissive bovine BLAC-20 and lapine EREp cells (13). Although this effect of bTat was described as being independent of bTAR, we wondered if the absence of appropriate RNA tethering of bTat could play some role in these cells.…”

“…The plasmid targets pHIVSCAT and pBLTRCAT have been described previously (13,14). To construct pHIVSCATbTAR, pHIVSCAT was cleaved at unique BglII and SacI sites, and bTAR sequence was inserted into pHIVSCAT by using two complementary bTAR oligonucleotides with BglII and SacI sites.…”

“…bTat transactivates strongly the BIV LTR (bLTR) in virally permissive canine (Cf2Th) cells and virally nonpermissive human (HeLa), monkey (CV1), and murine (3T3) cells (5,13). Surprisingly, bTat functions poorly and independently of bTAR in virally permissive bovine (BLAC-20) and lapine (EREp) cells (13).…”

Interactions between bovine retroviruses and the host immune system

“…The genome of BIV is the most complex of the nonprimate lentiviruses (Fong et al, 1995). In addition to the obligate retroviral structural genes gag, pol, and env, BIV contains at least five open reading frames (ORFs) in a "central region"…”