The Host Cell Sulfonation Pathway Contributes to Retroviral Infection at a Step Coincident with Provirus Establishment

Abstract

The early steps of retrovirus replication leading up to provirus establishment are highly dependent on cellular processes and represent a time when the virus is particularly vulnerable to antivirals and host defense mechanisms. However, the roles played by cellular factors are only partially understood. To identify cellular processes that participate in these critical steps, we employed a high volume screening of insertionally mutagenized somatic cells using a murine leukemia virus (MLV) vector. This approach identified a role for 3′-phosphoadenosine 5′-phosphosulfate synthase 1 (PAPSS1), one of two enzymes that synthesize PAPS, the high energy sulfate donor used in all sulfonation reactions catalyzed by cellular sulfotransferases. The role of the cellular sulfonation pathway was confirmed using chemical inhibitors of PAPS synthases and cellular sulfotransferases. The requirement for sulfonation was mapped to a stage during or shortly after MLV provirus establishment and influenced subsequent gene expression from the viral long terminal repeat (LTR) promoter. Infection of cells by an HIV vector was also shown to be highly dependent on the cellular sulfonation pathway. These studies have uncovered a heretofore unknown regulatory step of retroviral replication, have defined a new biological function for sulfonation in nuclear gene expression, and provide a potentially valuable new target for HIV/AIDS therapy.

Cellular Transcription Factor ZASC1 Regulates Murine Leukemia Virus Transcription

“…The viral genome plasmids pMMP-nls-LacZ, pCMMP-EGFP, pCMMP-SEAP-IRES-GFP, pCMMP-IRES-GFP, pCMMP-CD4-EGFP, pHIV-TVA800-hcRED, pRET, RCASBP(A)-AP, pLEGFP, pQCLIN, and pLenti6/V5-GW/lacZ have been described previously (5). The viral genome plasmid pCMMP-luciferase was generated by replacing the enhanced green fluorescent protein (EGFP) gene in pCMMP-EGFP with the firefly luciferase gene (fLuc) from pGL3-Basic (Promega, Madison, WI).…”

“…The source and growth conditions for Chinese hamster ovary cells (CHO-K1), CHO-TVA800, human embryonic kidney 293T cells, THP-1, and human Jurkat cells were described elsewhere (5,30). Mouse NIH 3T3 and EL4 cells were obtained from the ATCC (Manassas, VA) and cultured as suggested by the distributor.…”

“…GFP-ZASC1 expression was induced with 1 g/ml doxycycline. The procedures used to produce the retroviral vectors and titer of each viral stock are described in detail elsewhere (5).…”

“…However, it is likely that other important cellular factors remain to be identified. To illustrate this point, we recently used a forward genetic approach, based upon retroviral insertional mutagenesis in CHO-K1 cells, to uncover an unprecedented role for the host cell sulfonation pathway in regulating HIV-1 and MLV gene expression (5). The role of this cellular pathway in provirus transcription was not detected by multiple genome-wide RNA interference (RNAi)-based screens (4,18,34,37).…”

Evidence for association with hepatocellular carcinoma at the PAPSS1 locus on chromosome 4q25 in a family-based study

“…34,36 It was recently shown that PAPSS1 influenced retroviral replication by affecting a step during provirus establishment. 37 It is noted that multiple SNPs associated with HCC clustered within the PAPSS1 ATP sulfurylase domain that included exons 6 -12 and 3 0 -untranslated region. Two rare missense variants in this domain have been reported in other races; one of which can change in vitro substrate kinetic properties.…”