Although the frequency of neurological and neurobehavioral dysfunction in HIV-1-infected individuals has been reduced by HAART, nonetheless, the CNS may represent an important reservoir or sanctuary site for HIV-1 during virally-suppressive therapy.
In Specific Aim I, unique molecular mechanism(s) of HIV-1 persistence in astrocytes will be examined. We have recently demonstrated that HIV-1 Rev binds to, and functionally interacts with, a DEAD-box helicase, DDX1. This moiety binds to a specific motif in Rev (entitled NTS), which is involved with proper subcellular localization of Rev (i.e., nuclear greater than cytoplasmic). Recent data suggest that Rev is hyper-localized in primary astrocytes within the cytoplasm. Thus, we hypothesize that the DDX1 cellular co-factor may be involved with low-level residual HIV-1 replication in astrocytes. Presence and quantitation of DDX1 in human astrocytes, as well as potential co-localization with Rev, will be explored. Direct interactions between Rev plus selected Rev mutants and DDX1 will be assessed with a mammalian two-hybrid system. As well, over-expression of DDX1 and anti-sense and RNAi inhibition of DDX1 mRNA expression in astrocytes will be used to evaluate this potentially unique CNS-cell specific mechanism inducing HIV-1 persistence in human astrocytes.). A translation component will utilize single cell microdissection technology to directly probe the Rev:DDXl axis in vivo in the brains of diverse HIV-1-infected individuals.
In Specific Aim II, we will evaluate, in a complementary fashion with Specific Aim I, the potential additional and/or alternative mechanisms for alterations of Rev subcellular compartmentalization in low-level viral production in astrocytes. This will include analysis of potential splice variants of DDX1, and a hypothesized cellular inhibitor of DDX1 binding to NTS which is astrocyte-specific. As well, identification and purification of astrocyte-specific NIS binding cellular proteins which may alter subcellular compartmentalization will be analyzed by affinity chromatography and/or phage-peptide display libraries. Thus, these complementary approaches will be utilized to analyze, on molecular levels, the mechanisms of HIV-1 reservoirs and persistence in human astrocytes. ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
1R01MH074375-01A1
Application #
7005594
Study Section
NeuroAIDS and other End-Organ Diseases Study Section (NAED)
Program Officer
Joseph, Jeymohan
Project Start
2005-09-15
Project End
2009-08-31
Budget Start
2005-09-15
Budget End
2006-08-31
Support Year
1
Fiscal Year
2005
Total Cost
$370,105
Indirect Cost
Name
Thomas Jefferson University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
053284659
City
Philadelphia
State
PA
Country
United States
Zip Code
19107