The long-term goal of this proposal is to identify the pathophysiology underlying HIV associated COPD.
People living with HIV (PLWH) show increased incidence of COPD even when compensated for smoking status.
MicroRNAs manage the cellular transcriptome and play important roles in health and disease. We have shown
that HIV Tat and Transforming Growth Factor-beta (TGF-ß) signaling dysregulate the airway microRNAome.
Dysregulation of the microRNAome can effect multiple signaling pathways and this can affect cellular
homeostasis. COPD is a multifactorial pathology and involves dysregulation of diverse signaling pathways that
converge towards airway mitochondrial disfunction, circadian dysregulation, remodeling, inflammation, etc. TGF-
ß signaling is induced by both HIV Tat and cigarette smoke and this is significant since a significant proportion
of PLWH are addicted to nicotine and smoke tobacco/cigarettes, possibly exacerbating their progression to
COPD. Hence, Tat and TGF-ß-induced aberrant microRNAome and its effects on the airway
transcriptome can serve as an initiating events with downstream consequences on critical signaling
nodes and pathways. Identifying these pathways and these critical signaling nodes can help tailor interventions
to improve long-term pulmonary outcomes in PLWH.
Lungs are important HIV reservoirs and we, and others have shown that airway epithelial cells express
canonical HIV receptors and can be infected with HIV. A recent study shows that the bulk of HIV reservoirs are
transcriptionally active even with suppressive cART. Hence, silencing HIV transcription can decrease HIV
proteins like Tat in the airway. Gene editing using CRISPR/Cas9 has tremendous potential in eliminating HIV
reservoirs and reports have shown excision of HIV from infected cells. However, CRISPR based HIV excision is
vulnerable to viral escape as well as off-target effects. Cyclin T1, a component of P-TEFb, (heterodimer of
cyclinT1 and CDK9) is bound by HIV Tat and plays a pivotal role HIV transcription and its inhibition abolishes
HIV transcription. Hence CRISPR mediated CyclinT1 inactivation will completely block HIV transcription, locking
the HIV proviral DNA in an inactive form. We will use our patented Pol II HIV LTR-drosophila hsp70 fusion
monopromoter to co-express CyclinT1 gRNA and Cas9 (and effect CyclinT1 inactivation) only in HIV
infected cells. Expression is self-limiting as our fusion promoter also requires CyclinT1 thereby minimizing off-
target effects. Aim 1 will determine the mechanism by which HIV Tat and TGF-ß dysregulate the airway
microRNAome and its effects on the transcriptome to identify signaling pathways involved in HIV associated
COPD. Aim 2 will use an inducible CRISPR syste to inactivate CyclinT1 only in HIV infected cells and lock the
proviral DNA in a transcriptionally inactive state. Understanding the signaling pathways that promote HIV-
associated COPD and silencing HIV transcription in reservoirs will prevent lung function decline and
development of COPD in PLWH.