Presenting Author:
Theron Russell, M.S.
Principal Investigator:
Peter Penzes, Ph.D.
Department:
Physiology
Keywords:
schizophrenia, single nucleotide variant, guanine nucleotide exchange factor, small GTPase, cerebral cortex, neuron, gen... [Read full text]
Location:
Third Floor, Feinberg Pavilion, Northwestern Memorial Hospital
B165 - Basic Science
Deleterious effects of a schizophrenia-associated nonsynonymous exonic KALRN variant
Large scale genetic studies have revealed that rare sequence variants, such as copy number variations and single nucleotide polymorphisms (SNPs), in glutamatergic synaptic plasticity genes are enriched in subjects with schizophrenia (SZ). The majority of SZ-associated SNPs are located in intronic regions, leading to dysregulated gene expression rather than altered protein structure and function. Nevertheless, several exonic SZ-associated SNPs have been identified in both GWAS and resequencing studies, although their functional effects have yet to be examined. One such SNP, KALRN:p.P2255T, displays relatively high penetrance (OR ≈ 2) and has been predicted to be damaging in silico. Therefore, we sought to characterize the effects of this SNP in the context of the dual Rac1 and RhoA guanine nucleotide exchange factor Kalirin-9 (Kal9) protein. Here we show that overexpression of Kal9-P2255T leads to diminished basal dendritic branching and reduced dendritic spine area and breadth in primary cortical pyramidal neurons, as compared with overexpression of wildtype Kal9. The small GTPases Rac1 and RhoA are known to effect cytoskeletal changes that underlie spine and dendritic dynamics, so we tested the effects of Kal9-P2255T on their activity, and determined that RhoA activation is increased in heterologous cells expressing the SNP rather than wildtype Kal9, whereas Rac1 activation is unchanged. Because elevated levels of Kal9 are seen in subjects with SZ, we conjectured that another possible mechanism by which Kal9-P2255T exerts its effects is by increasing Kal9 levels. Following transfection of both heterologous cells and primary cortical neurons with equal amounts of Kal9 cDNA, we found that Kal9-P2255T protein levels were higher than those of wildtype. Pharmacological inhibition of protein translation followed by analysis of the rate of Kal9 degradation indicated that the P2255T variant does not confer changes to protein stability. However, inhibition of transcription allowed us to establish that the increased Kal9-P2255T protein levels are due to the SNP imparting increased stability to Kal9 mRNA. Taken together, the data suggest that both heightened Kal9-P2255T RhoA-GEF catalytic function and increased Kal9-P2255T mRNA expression combine to give rise to the cortical ultrastructural pathology that is thought to bring about SZ symptomatology.
