Replication of HIV-1 viruses in the presence of the Portland alpha1-antitrypsin variant (alpha1-PDX) inhibitor.
The Portland alpha1-antitrypsin variant (alpha1-PDX) inhibits gp160 cleavage into gp120 and gp41 by different prohormone convertases (PCs) including furin, PC5 and PC7. Jurkat cells stably transfected with this inhibitor (J-PDX cells) and, as controls, Jurkat cells transfected with the empty vector (J-pcDNA3) were tested for their susceptibility to HIV-1 infection. We found that HIV-1 replication was significantly impaired in J-PDX cells. However, the analysis of the infectivity of HIV-1 viruses produced in J-PDX cells on different days during the infection indicated that they recovered infectivity starting from 13 days post-infection. The sequencing of viruses collected earlier and later from J-PDX cells revealed no mutations in envelope-glycoprotein precursor (Env) maturation sites or in the N-terminal sequence of gp41 fusion peptide, which plays a key role in membrane fusion. Although conserved mutations were detected at the C-terminus of the gp41 fusion peptide and ectodomain, the replication of mutant HIV-1 viruses produced on day 20 in J-PDX cells was inhibited at a similar level to wild-type viruses after a second passage in J-PDX cells. We then investigated the expression of the alpha1-PDX protein, and found that HIV-1 replication activated its proteolysis since the 54 kDa cleaved form became predominant later on in the infection. In contrast, the expression of PC7, a protein that is transported through the secretory pathway, was unaltered in HIV-1 infected cells. We conclude that recovered HIV-1 infectivity in J-PDX cells was due to a loss of alpha1-PDX activity via its extensive processing by intracellular proteases that cleave it through the substrate pathway.
Bahbouhi
B
B
Laboratoire d'immuno-virologie, EA 30-38 Université Paul Sabatier, UFR/SVT, 31062 Toulouse, France.
Seidah
N G
NG
Bahraoui
E
E
D015153:Blotting, Western; D015496:CD4-Positive T-Lymphocytes; D017124:Conserved Sequence; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D019302:HIV Envelope Protein gp160 / Q000378:metabolism; D015700:HIV Envelope Protein gp41 / Q000378:metabolism; D015497:HIV-1 / Q000502:physiology*; D006801:Humans; D019169:Jurkat Cells; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014162:Transfection; D014779:Virus Replication; D000515:alpha 1-Antitrypsin / Q000494:pharmacology*
D015699:HIV Envelope Protein gp120; D019302:HIV Envelope Protein gp160; D015700:HIV Envelope Protein gp41; D010455:Peptides; D011993:Recombinant Fusion Proteins; D000515:alpha 1-Antitrypsin; C113421:alpha 1-antitrypsin Portland