In silico characterization and analysis of transcriptional inhibition of PDGF in glioblastoma tumor cells by CRISPR-C13a technique

Glioblastoma, also known as glioblastoma multiforme (GBM), is the most aggressive cancer that begins withinthe brain. Initially, signs and symptoms of glioblastoma are non-specific .The cause of most cases isunclear. Uncommon risk factors include genetic disorders such as neurofibromatosis and Li–Fraumeni syndrome.Platelet-derived growth factor (PDGF) is one of numerous growth factors that regulate cell growth and division.In particular, PDGF plays a significant role in angiogenesis, the growth of blood vessels from already-existingblood vessel tissue, mitogenesis, i.e. proliferation, of mesenchymal cells such as fibroblasts, osteoblasts,tenocytes, vascular smooth muscle cells and mesenchymal stem cells. PDGFA and PDGFα receptors are coexpressedin glioblastoma and oligodendroglioma. The PDGFRα gene is amplified, mutated, or rearranged in afraction of glioblastoma tumors. Genetic aberrations and overactivity of the PDGFα receptor signaling pathwayare important events in the development of a subset of glioblastomas. RNA targeting was a challenging issue ingenetic engineering. Even CRISPR-Cas with its great applications, cannot target RNA. Recently, researchershave found out that a C13a protein in Leptotrichia shahii bacteria has two RNase activities, one for cutting itsRNA target and the other for processing the CRISPR RNA (crRNA). Functional studies revealed that C13a is aRNA-guided RNA-targeting CRISPR effector. We design crRNA by CRISPR-RT for targeting PDGF RNA.After designing crRNA that targets the PDGF RNA, its specificity and structure was evaluated. The best crRNAwas selected, it could target PDGF mRNA very specific and reduce its RNA level.

Keywords:

CRISPR-C13a , Glioblastoma , PDGF , crRNA , CRISPR-RT