We discovered that a completely uncharacterized long noncoding RNA (lncRNA) ADIRF- AS1 binds to the tumor suppressor Polybromo- and BRG1-associated factors-containing (PBAF) chromatin modifying complex, including the three unique protein components of the PBAF complex: ARID2, BRD7, and PBRM1. The PBAF complex bromodomain containing protein PBRM1 is mutated in approximately 40% of clear cell renal carcinoma (ccRCC), where loss of PBRM1 increases proliferation and overall tumor burden. Publicly available patient data revealed that ADIRF-AS1 is more highly expressed in ccRCC tumors compared to normal tissues. We overexpressed ADIRF-AS1 in a panel of ccRCC cell lines and found that ADIRF-AS1 overexpression significantly promoted proliferation only in ccRCC cell lines with wildtype expression of PBAF protein components. Importantly, we found that overexpression of ADIRF-AS1 decreased protein expression of PBAF protein components. Next, we sought to identify how ADIRF-AS1 negatively regulates the PBAF complex. We performed a lncRNA pulldown assay followed by proteomics analysis and found that along with components of the PBAF complex, ADIRF-AS1 also associates with an E3 ubiquitin ligase TRIM25, which has been shown to target PBRM1 for proteasomal degradation. We validated that silencing TRIM25 increases PBRM1 expression and found that proteasome inhibitors rescued decreased expression of PBRM1 after ADIRF-AS1 overexpression. Therefore, I hypothesize that ADIRF-AS1 interacts with the PBAF complex to promote its degradation by acting as a scaffold for PBRM1 and TRIM25, promoting tumor formation and progression. To address this hypothesis, we propose two aims to: (1) Determine the role of ADIRF-AS1 expression in PBAF mediated metabolism and tumorigenesis, and (2) Characterize the mechanism by which ADIRF-AS1 negatively regulates the PBAF complex through TRIM25 to promote tumorigenesis.
Many patients with kidney cancer (clear cell renal carcinoma, ccRCC) lose the tumor suppressor PBRM1, and loss of PBRM1 has been shown to accelerate tumor growth. We discovered that a completely uncharacterized noncoding RNA, ADIRF-AS1, binds to both PBRM1 and TRIM25 that is known to target PBRM1 for degradation; and, loss of ADIRF-AS1 increases PBRM1 expression, while increased ADIRF-AS1 expression decreases PBRM1 in ccRCC cell lines. As ADIRF-AS1 is upregulated in ccRCC tumors compared to normal tissue, understanding how ADIRF-AS1 functions in kidney cancer may lead to new therapeutic strategies.
