The piR-31115-PIWIL4 complex promotes the migration of the triple-negative breast cancer cell lineMDA-MB-231 by suppressing HSP90AA1 degradation

Abstract

Background

P-element-induced wimpy testis (PIWI) proteins bind to PIWI-interactingRNAs (piRNAs) to form the piRNA/PIWI complex, which affects protein regulation. PIWIL4, a member of the PIWI family, has been demonstrated in recent studies to promote the migration of triple-negative breast cancer (TNBC) cell line MDA-MB-231. However, the molecular mechanisms underlying cell migration remain obscure.

Methods

RNA immunoprecipitation and real-time PCR assays were conducted to detect piRNAs binding to PIWIL4. piRNA mimics and inhibitors were employed to modify piRNA expression in MDA-MB-231 cells. Cell migration assays were carried out using transwell inserts. Co-immunoprecipitation (co-IP) combined with mass spectrometry (MS) was performed to identify the proteins that interacted with PIWIL4 under the regulation of piRNA. Western blotting (WB) was utilised to detect the regulatory relationship between the piRNA/PIWIL4 complexes and the mutually-binding proteins.

Results

RNA Immunoprecipitation (RIP) results revealed that PIWIL4 bound to piR-31115 in the MDA-MB-231 cells. Transwell assays demonstrated that piR-31115 promoted the migration of MDA-MB-231 cells via PIWIL4. Co-IP coupled with MS results showed that piR-31115 promoted the binding of PIWIL4 to HSP90AA1 in MDA-MB-231 cells, and this interaction protected HSP90AA1 from degradation. Knockdown of HSP90AA1 in MDA-MB-231 cells attenuated the promoting effects of piR-31115/PIWIL4 on cell migration.

Conclusions

Our findings cast light on a novel molecular pathway through which piR-31115 promotes the migration of MDA-MB-231 TNBC cells by regulating the interaction between PIWIL4 and HSP90AA1.