Fig 2: PAK1 Expression in schizophrenia. A) Detection of PAK1 in human gray matter extracts. Bands corresponding to the predicted molecular weights of PAK1 and Tubulin are readily detected in human gray matter extracts from subjects with schizophrenia (S) and matched control subjects (C). MW, Molecular Weight Markers. B) Comparison of PAK1 expression (normalized to tubulin expression) for the 25 pairs of subjects. Each point represents a pair of subjects. The diagonal reference line represents a control: schizophrenia ratio of one. Points above the line indicate increased expression in schizophrenia subjects, points below the line indicate decreased expression in schizophrenia subjects. C) Mean (SD) expression PAK1:tubulin in schizophrenia and control subjects. D) Correlation of within pair differences in log PAK1 expression with the corresponding measure of log total kalirin protein expression (r = 0.55, p = 0.004). E) Correlation of within pair differences in PAK1 expression with the corresponding measure of spinophilin protein expression (r = 0.61, p = 0.004).

Fig 3: Effects of postmortem interval on PAK1 expression. A) Example western blot of PAK1 and Tubulin in mice in which the interval from sacrifice to brain harvesting was experimentally varied between 0 and 48 hours. MW, Molecular Weight Markers. B) Optical densities for PAK1 and Tubulin in mice in which the interval from sacrifice to brain harvesting was experimentally varied between 0 and 48 hours. Two sets of mice (each set containing one mouse for each PMI) were tested. Sets were run concurrently on separate gels, and the run was repeated. Mean (SEM) values for the four observations at each time point are shown. Time points sharing the same superscript letter differ significantly.

Fig 4: PAK1 Signaling Pathway.PAK1 resides in inactive homo- or heterodimers. Binding of the Rho-GTPase, RAC1 (or CDC42) causes dissociation of the dimers and activation of PAK1. PAK1 activates LIMK1, which inhibits cofilin-mediated f-actin depolymerization. PAK1 may be subject to activation by non-GTPase mechanisms, and can activate other effector pathways, some of which (e.g. MLCK and MLC) may also impact spine dynamics. Blue indicates promotion of dendritic spine persistence, red indicates promotion of spine elimination.

Fig 5: Processing of human tissue for PAK1. A–C) show an example of a frozen coronal slab through the right temporal lobe from which a block containing the STG and Heschl's gyrus (located between the arrows in C) has been excised and mounted for cryostat sectioning. The boundary between the gray and white matter of Heschl's gyrus (dotted line) was undercut so that only gray matter was collected for protein extraction. D) Demonstrates our approach to quantification of PAK1 and tubulin. Lanes from all blots were simultaneously imported into the analysis software, and for quantification of each protein the peaks were aligned, and then a single band definition applied to all blots concurrently, indicated by the black horizontal lines. E) PAK1 optical density as a function of micrograms of protein loaded per lane. The mean (SD) of 3 human subjects, assayed together in duplicate runs is shown. It can be seen that the protein loading used in the comparison of schizophrenia and control subjects (20 µg) sits within the linear detection range. F) Western blot demonstrating isoform specificity of anti-PAK1 antibody. Lanes were loaded with 0.5 µg of each full length GST-tagged recombinant protein (ProQinase # 0357-0000-1, 0304-0000-1, 0422-0000-1, Freiburg, Germany), and detected with Coomassie Blue (top) or with anti-PAK1 antibody (bottom) as described for human and mouse studies.