Genetic Suppression of SMN Mutations in Spinal Muscular Atrophy

Spinal Muscular Atrophy is a devastating neuromuscular disease caused by insufficient amounts of SMN protein.
SMA is caused by loss or mutation of the SMN1 gene and retention of the SMN2 gene. The SMN2 gene is a
modifier of phenotype where milder SMA cases having more copies of SMN2. Rarely SMA patients have a
missense mutation in the SMN1 gene. We can use these mutations and the protein domains they disrupt to
study the function of the SMN protein. We have shown that SMA missense mutations are not functional by
themselves but can function in the presence of some full-length wild-type SMN protein. Furthermore, we have
shown that N and C-terminal SMN missense mutations can complement each other and rescue snRNP assembly
in the complete absence of full-length wild-type SMN protein in mice. We have developed cell line that
conditionally removes functional SMN to allow us test SMN missense mutations in culture. We have also used
this cell line to test for suppressors of the SMNE134K mutation. We have identified a suppressor in the SmF
protein that fully restores snRNP assembly lost due to the SMN E134K mutation. We now have a system to
screen for suppressors of SMN missense mutations. In this proposal we will test the SmF suppressor we have
found in two different SMN E134K mouse models and determine if this mutation rescues the SMA phenotype
and survival of the SMA mouse. Thus, we can study the separate functions of SMN in snRNP assembly from
the function of SMN in the axon. We will screen for additional suppressors using other SMN patient derived
mutations to test other functional domains of SMN. We will investigate the role of SMN in the axon independent
of Sm assembly by introducing HuD and truncated forms of SMN into the SMA mice via scAAV9. We have
shown in cells that Smn exon2B is not required for cell survival. We will test scAAV9-Smn∆2 in SMA mice to
confirm this finding and rescue the SMA phenotype. Finally we will test the role of profilin in axonal function in
the SMA mouse using the SMNS230L mutation. Using genetic mutations we can dissect the functions of SMN
in splicing and in the axon to resolve the underlying mechanism by which reduced SMN protein causes SMA.

Grant Number: 5R01NS123736-05
NIH Institute/Center: NIH
Principal Investigator: ARTHUR BURGHES