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. 2024 May 3;300(6):107342. doi: 10.1016/j.jbc.2024.107342

Figure 7.

Figure 7

K-27 is critical for Hsp82-dependent stability and activity of Rad51.A, Western blot showing Rad51 level in whole cell extracts of WT, K27Qhsp82, and K27Rhsp82 strains. B, the above experiment was repeated and the quantification of Western blots showed a four-fold reduced stability of Rad51 in K27Qhsp82 strains; the mean value (±SD) was plotted; p values were calculated using the two-tailed Student’s t test (∗∗p < 0.01). C, Western blot showing Rad51 level in whole cell extracts of WT, K274Qhsp82, and K274Rhsp82 strains. D, the quantification of independent repeats of the above experiment showed increased stability of Rad51 in both K274Qhsp82 and K274Rhsp82 strain; the mean value (±SD) was plotted; p values were calculated using the two-tailed Student’s t test (∗∗p < 0.01, ∗p < 0.05). E, Western blot showing Rad51 level in whole cell extracts of WT, K27QK274Qhsp82, and K27RK274Rhsp82 strains. F, the above experiment was repeated and the quantification of Western blots depicted a two-fold reduced stability of Rad51 in K27QK274Qhsp82 strain, whereas that remained unchanged in K27RK274Rhsp82 strain; the mean value (±SD) was plotted; p values were calculated using the two-tailed Student’s t test (∗∗∗∗p < 0.0001, NS, not significant). G, Rad51 levels in K27Qhsp82 (upper panel), K27QK274Qhsp82 (middle panel), and WT strains (bottom panel) were monitored in presence of MG132. H, schematic representation of the gene targeting cassette consisting the upstream and downstream homologous stretches of ADH4 gene, flanking the ADE2 gene, and a KANMX selectable marker. Random integration generates Ade+G418SR colonies, while targeted integration results in Ade+G418SS colonies. I, gene targeting assay showed 30% and 40% reduced gene targeting efficiency in K27Qhsp82 and K27QK274Qhsp82 strains, respectively, whereas there was no change in gene targeting efficiency in other hsp82 mutant strains; the mean value (±SD) was plotted; p values were calculated using the two-tailed Student’s t test (∗∗p < 0.01, ∗p < 0.05, NS, not significant).