Abstract 3100: BRCA1-RpL17 Interaction Regulates Angiogenesis

Abstract

Background: Angiogenesis, forming new blood vessels from existing ones, is crucial in development, diabetes, and cancer. We found that Large Ribosomal subunit protein 17 (RpL17) played a key role in endothelial cell (EC) proliferation and angiogenesis. Reduced RpL17 (KD) promoted angiogenesis in human ECs and RpL17-deficient mouse retinas. RNA-sequencing and bioinformatics suggested a role for Early growth response 1(EGR1) transcription factor. Bioinformatics revealed Breast Cancer gene 1 (BRCA1) as a potential upstream regulator of RpL17, suggesting a novel pathway for angiogenesis. Hypothesis: BRCA1 interacts with RpL17 and upregulates EGR1 to stimulate vessel elongation in angiogenesis. Goal: Determine the role of BRCA1-RpL17 interaction in angiogenesis and their potential feedback loop via EGR1. Methods/Approach: RNA-sequencing of mouse ECs, whole retina mount staining (comparing wild-type and RpL17+/- mice). Tube formation assays with HUVECs to investigate angiogenesis after RpL17, BRCA1, and EGR1 knockdowns. Bioinformatics analysis with Python for RNA-seq data. Immunoblotting and RT-qPCR to measure protein and DNA expression. Immunofluorescence (IF) staining of HUVECs to observe BRCA1, RpL17, and EGR1 expression and localization. Preliminary Results: Whole retina mounts showed increased angiogenesis in RpL17-deficient mouse retinas compared to wildtype (p<0.009). HUVECs with RpL17 KD had increased junctions and longer branches than control HUVECs (p<0.0002). RNA-seq identified increased EGR1 expression (q<2.5x10 -8 ) in RpL17 KD ECs, confirmed with RT-qPCR (p<0.002). Bioinformatics predicted BRCA1 as a regulator of RpL17. HUVEC IF staining showed decreased RpL17 expression after BRCA1 KD. Angiogenesis assays of BRCA1 KD ECs exhibited an angiogenic phenotype like RpL17 KD, rescued following double BRCA1/RpL17 KD. Conclusion: BRCA1-RpL17 interaction stimulates angiogenesis and may be regulated by an EGR1 feedback loop, likely by EC proliferation. Detailed understanding of this pathway will provide new therapies to regulate angiogenesis.