(A) Using denatured IP, Munc18-1 was pulled-down from cell lysate of HEK293T cells expressing M18WT or M18Y473A together with SFK kinases or an empty vector as control. Munc18-1 was then immunoblotted for tyrosine phosphorylation using the 4G10 antibody. The total amount of Munc18-1 was detected after stripping and reblotting for Munc18-1. 