The aim of the project is to explain the molecular mechanisms underlying the neurological symptoms that patients with specific mutations for Slk and Ola1 mutations have (causing complete knockdown of protein expression), such as intellectual disability and sensory phenotypes. This project consists of a collaboration between my PI Dr. Leena Ibrahim and the Professor of Human Genetics at the Alfaisal University, Dr. Fowzan Alkuraya.
For each of the three mutations we have designed different experimental approaches. In order to simulate slk and ola1 mutations we are implementing CRISPR-Cas9 gene editing technologies in vitro using mouse cortical neuronal cultures. Similarly, to simulate these mutations in neurons, we received fibroblast samples from the affected clinical patients and with the use of a novel technology consisting of direct transdifferentiation from fibroblasts into neurons we will be able to convert patient s fibroblast cell lines and analyze them, morphologically and functionally.
After successfully creating the mutated neuronal cell lines, our goal is to be able to analyze the functionality of different proteins that might be related to the SLK and OLA1. This will be done by the analysis of protein colocalization using immunocytochemistry and high-resolution bioimaging techniques. Together with this, we aim to investigate the physiological significance of these mutations at morphological and functional level. For this purpose we will be performing different tests to evaluate synaptic alterations happening in the knockdown and mutated cell lines using patch-clamp recordings.
Up to date there is no information regarding the effects that OLA1 might have in the context of neurodevelopment. However, our recent findings point out that the protein could have a regulatory role through the modulation of pathways involved neurite outgrowth and migration due to its binding activity with HSP70 providing stability and longer lasting activity to this protein that has proven to be intrinsically related to neurodevelopmental processes.