Fig 1. Mass spectrometry identification of actin as a binding partner of RACK1.

A SH-SY5Y cells were treated with 10 μM FSK for 30 min, RACK1 was IPed from cell lysate and proteins were resolved by SDS-PAGE which was stained with Deep Purple™ to visualize proteins. Gel slices from both RACK1 and control IgG IPs were digested in-gel and the resulting peptides were submitted to tandem mass spectrometry (MS/MS) sequencing for protein identification, n = 2. B Identification of actin as a putative RACK1 binding protein. β-actin (ACTB), γ-actin (ACTG), alpha skeletal muscle actin (ACTS), aortic smooth muscle actin (ACTA), alpha cardiac muscle actin (ACTC) were identified in the RACK1 IP. A sum of score of peptides that were matched to the best hit within the family of homologous protein sequences (¹). Calculation of the percent sequence coverage included all peptides matched to the protein, i.e., both unique and common peptides (²). Differentiation between ACTC, ACTS and ACTA proteins was not possible on the basis of the data since the identified peptides were common to all of them (³). Differentiation between β-actin and γ-actin proteins was not possible on the basis of the data since the identified peptides were common to both isoforms (⁴).