Reticulocalbin 3 Is a Novel Mediator of Glioblastoma Progression

SIMPLE SUMMARY: Glioblastoma is the deadliest primary brain tumor. Current treatment strategies for glioblastoma are not effective, and patients exhibit poor survival rates. Emerging studies implicated glioma stem cells (GSCs) in tumor initiation, progression, and therapy resistance. There is an unmet need to identify new therapeutic candidates for glioblastoma. Our study showed that reticulocalbin 3 (RCN3), an ER lumen residing Ca(2+) binding protein, is overexpressed in glioblastoma and associated with poor survival rates. The reduction in RCN3 expression using shRNA or gRNA resulted in reduced proliferation and self-renewal of GSCs. The RNA-seq results showed that RCN3 knockdown altered the expression of several genes related to translation, ribosome, stem cell differentiation, and extracellular matrix. In silico analysis of glioblastoma patient datasets demonstrated a positive correlation of RCN3 with ribosomal pathway genes. Importantly knockdown of RCN3 significantly enhanced the survival of tumor-bearing mice. Our study suggests that RCN3 could be a potential therapeutic target in glioblastoma. ABSTRACT: Glioblastoma is the most common malignant primary brain tumor. Molecular mechanisms underlying the pathobiology of glioblastoma are incompletely understood, emphasizing an unmet need for the identification of new therapeutic candidates. Reticulocalbin 3 (RCN3), an ER lumen-residing Ca(2+) binding protein, plays an essential role in protein biosynthesis processes via the secretory pathway. Emerging studies demonstrated that RCN3 is a target for therapeutic intervention in various diseases. However, a knowledge gap exists about whether RCN3 plays a role in glioblastoma. Publicly available datasets suggest RCN3 is overexpressed in glioblastoma and portends poor survival rates. The knockdown or knockout of RCN3 using shRNA or CRISPR/Cas9 gRNA, respectively, significantly reduced proliferation, neurosphere formation, and self-renewal of GSCs. The RNA-seq studies showed downregulation of genes related to translation, ribosome, and cytokine signaling and upregulation of genes related to immune response, stem cell differentiation, and extracellular matrix (ECM) in RCN3 knockdown cells. Mechanistic studies using qRT-PCR showed decreased expression of ribosomal and increased expression of ER stress genes. Further, in silico analysis of glioblastoma patient datasets showed RCN3 expression correlated with the ribosome, ECM, and immune response pathway genes. Importantly, the knockdown of RCN3 using shRNA significantly enhanced the survival of tumor-bearing mice in orthotopic glioblastoma models. Our study suggests that RCN3 could be a potential target for the development of a therapeutic intervention in glioblastoma.