Aims:
Breast cancer is a complex disease due to the high heterogeneity within the tumour microenvironment, especially in the triple negative breast cancer (TNBC) subtype. BRCA1 serves as one of the most common breast cancer markers with its essential functions ranging from DNA damage repair to cellular homeostasis. Cavin3 has been linked to breast cancer and shown to associate with BRCA1, a key player in DNA damage repair and cellular homeostasis. Therefore, we would like to develop a system to characterise the interaction between Cavin3 and BRCA1 with the goal of identifying the specific domains in both proteins responsible for this binding. Then, we characterise other components required for the BRCA1-Cavin3 interplay. Finally, we examine the consequences of this interaction in human breast cancer patient samples. While caveolae have been previously linked to breast cancer, a comprehensive study examining the expression levels of caveola-associated proteins in breast cancer is still lacking. As part of this aim, we sought to generate a transcriptomic profile of all caveola-associated proteins across four breast cancer subtypes.
Methods:
HeLa WT and Cavin3-null cell lines were used to perform GFP-nanotrap co-immunoprecipitation, and western blotting. Single cell RNA sequencing analysis (Seurat package) was applied to determine the impact of BRCA1-Cavin3 interaction in the clinical breast cancer samples.
Results:
Cavin3 interacts with and is stabilised by the first 100 amino acids, encompass RING domain, of BRCA1 through its first helical region (HR1). Interestingly, Cavin3 maintains its interaction with different pathogenic BRCA1 mutations. Furthermore, our transcriptomic analysis of breast cancer samples indicates that Cavin3 and BRCA1 may be functionally correlated in spindle formation, cell cycle checkpoint, and DNA damage repair by co-regulating the gene expression of BRCA1 downstream targets. The expression levels of both CAVEOLIN1 (CAV1) and CAVIN1 are significantly increased in cancer epithelial cells within the TNBC subtype. Similar to CAV1 and CAVIN1, CAV2 and CAVIN3 also showed increased expression levels in in cancer epithelial cells within the TNBC subtype, while CAV3, CAVIN2 and CAVIN4 were barely expressed in these cells.
Conclusions:
We successfully developed a system to characterise the BRCA1-Cavin3 interaction and other proteins might be associated with this complex. BRCA1-CAVIN3 transcriptional co-expression tends to be appeared in more aggressive and advanced breast cancer. In TNBC samples, CAV1 and CAVIN1 is significant upregulated while CAV2 and CAVIN3 shows increased expression level.