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. 2011 Sep 15;124(18):3164–3173. doi: 10.1242/jcs.087320

Fig. 3.

Fig. 3.

Rescue of st601 and e286 unc-45 mutants by transgenic UCS-containing proteins. (A) The UCS domain and TPR(−) construct are sufficient for significant rescue of st601 lethality. The percentage of dead eggs of the total progeny was measured after injection of the parental hermaphrodite with the specific constructs. Error bars indicate mean ± s.d. *P≤0.05; ***P<0.001. N. S., not significant. (B) The UCS domain and TPR(−) construct are sufficient to significantly rescue e286 motility at 25°C. Error bars indicate mean ± s.d. ***P<0.001. (C) The UCS domain and TPR(−) construct rescue A-band assembly in e286 at 25°C. The outline of worm muscle cell was faint in the UCS(−) or parental e286 mutant lines. (D) Quantification of A-band assembly rescue of transgenic e286 at 25°C. Error bars indicate mean ±s.d. ***P<0.001. (E) The UCS domain is expressed at lower levels compared with the other transgenic proteins, and the myosin-binding truncates rescue body-wall muscle myosin accumulation as does FL in e286. Transgenic expression of UNC-45 fragments was detected by immunoblots with anti-FLAG antibody. UCS band was quite weak with anti-FLAG antibody staining. Body-wall muscle-specific myosin heavy chains A and B were detected and quantified by immunoblots. Pharyngeal myosin heavy chain D was used as loading control. (F) The detection of UNC-45 truncates in e286 by rabbit polyclonal anti-C. elegans UNC-45 antibody. UCS and other fragments were detectable on the blots. Endogenous UNC-45 and FLAG-tagged full-length UNC-45 overlapped on the top of the immunoblot. Pharyngeal myosin heavy chain D was used as loading control. (G) A titration experiment of transgenic UCS worms in e286 was performed to semi-quantitatively compare the level of UCS expression with that of FL in FL transgenic e286 by reaction with rabbit polyclonal anti-C. elegans UNC-45 antibody. Endogenous UNC-45 and FLAG-tagged full-length UNC-45 overlapped on the top of the immunoblot. UCS was visible by polyclonal anti-C. elegans UNC-45 antibody. Pharyngeal myosin heavy chain D was used as loading control. (H) Table showing the ratios of protein amounts, body bends and A-bands between transgenic FL and UCS e286.