Plasma Proteomic Profiling in HIV-1 Infected Methamphetamine Abusers

We wanted to determine whether methamphetamine use affects a subset of plasma proteins in HIV-infected persons. Plasma samples from two visits were identified for subjects from four groups: HIV+, ongoing, persistent METH use; HIV+, short-term METH abstinent; HIV+, long term METH abstinence; HIV negative, no history of METH use. Among 390 proteins identified, 28 showed significant changes in expression in the HIV+/persistent METH+ group over the two visits, which were not attributable to HIV itself. These proteins were involved in complement, coagulation pathways and oxidative stress. Continuous METH use is an unstable condition, altering levels of a number of plasma proteins

Proteomic Analysis of Early HIV‑1 Nucleoprotein Complexes

After entry into the cell, the early steps of the human immunodeficiency virus type 1 (HIV-1) replication cycle are mediated by two functionally distinct nucleoprotein complexes, the reverse transcription complex (RTC) and preintegration complex (PIC). These two unique viral complexes are responsible for the conversion of the single-stranded RNA genome into double-stranded DNA, transport of the DNA into the nucleus, and integration of the viral DNA into the host cell chromosome. Prior biochemical analyses suggest that these complexes are large and contain multiple undiscovered host cell factors. In this study, functional HIV-1 RTCs and PICs were partially purified by velocity gradient centrifugation and fractionation, concentrated, trypsin digested, and analyzed by LC–MS/MS. A total of seven parallel infected and control biological replicates were completed. Database searches were performed with Proteome Discoverer and a comparison of the HIV-1 samples to parallel uninfected control samples was used to identify unique cellular factors. The analysis produced a total data set of 11055 proteins. Several previously characterized HIV-1 factors were identified, including XRCC6, TFRC, and HSP70. The presence of XRCC6 was confirmed in infected fractions and shown to be associated with HIV-1 DNA by immunoprecipitation-PCR experiments. Overall, the analysis identified 94 proteins unique in the infected fractions and 121 proteins unique to the control fractions with ≥2 protein assignments. An additional 54 and 52 were classified as enriched in the infected and control samples, respectively, based on a 3-fold difference in total Proteome Discoverer probability score. The differential expression of several candidate proteins was validated by Western blot analysis. This study contributes additional novel candidate proteins to the growing published bioinformatic data sets of proteins that contribute to HIV-1 replication

Summary of differentially expressed proteins with statistical significance.

<p>*The protein sequences have no homolog sequence in UniProtKB database.</p

Changes in plasma protein expression levels between visits in all groups.^§

<p>NS: p-value is not significant.</p>§<p>- Changes are relative increases or decreases from the second visit compared to the first expressed as averaged for each group.</p

Subject demographic, psychiatric, and clinical characteristics.

<p><u>Abbreviations:</u> CART = combination antiretroviral therapy; SD = standard deviation; VL = viral load log10 copies/mL; IQR = interquartile range.</p>a<p>METH use disorder defined as meeting DSM-IV criteria for METH abuse or dependence.</p>b<p>These variables are restricted to METH+ subjects.</p>c<p>These variables are restricted to HIV+ subjects.</p

iTRAQ tag assignation for 4 biological replicates in experimental design.

<p>*P = Patient; G = Group; V = Visit.</p

Distribution of the Log₂ fold changes in proteins' expression in all groups.

<p>It is expected that proteins will be distributed around log2 of 0 ( = 1, no change between visits). We observed two additional peaks in Group 1 indicated by arrowheads; larger peak with log₂ of 1 corresponding to 2-fold increase and smaller peak with log₂ of approximately -2 corresponding to 7-fold decrease.</p