TY - JOUR
T1 - Effects of human deafness γ-actin mutations (DFNA20/26) on actin function
AU - Bryan, Keith E.
AU - Wen, Kuo Kuang
AU - Zhu, Mei
AU - Rendtorff, Nanna Dahl
AU - Feldkamp, Michael
AU - Tranebjaerg, Lisbeth
AU - Friderici, Karen H.
AU - Rubenstein, Peter A.
PY - 2006/7/21
Y1 - 2006/7/21
N2 - Six point mutations in non-muscle γ-actin at the DFNA20/26 locus cause autosomal dominant nonsyndromic hearing loss. The molecular basis for the hearing loss is unknown. We have engineered each γ-actin mutation into yeast actin to investigate the effects of these mutations on actin function in vivo and in vitro. Cells expressing each of the mutant actins as the sole actin in the cell were viable. Four of the six mutant strains exhibited significant growth deficiencies in complete medium and an inability to grow on glycerol as the sole carbon source, implying a mitochondrial defect(s). These four strains exhibited abnormal mitochondrial morphology, although the mtDNA was retained. All of the mutant cells exhibited an abnormally high percentage of fragmented/non-polarized actin cables or randomly distributed actin patches. Five of the six mutants displayed strain-specific vacuole morphological abnormalities. Two of the purified mutant actins exhibited decreased thermal stability and increased rates of nucleotide exchange, indicative of increased protein flexibility. V370A actin alone polymerized abnormally. It aggregated in low ionic strength buffer and polymerized faster than wild-type actin, probably in part because of enhanced nucleation. Mixtures of wild-type and V370A actins displayed kinetic properties in proportion to the mole fraction of each actin in the mixture. No dominant effect of the mutant actin was observed. Our results suggest that a major factor in the deafness caused by these mutations is an altered ability of the actin filaments to be properly regulated by actin-binding proteins rather than an inability to polymerize.
AB - Six point mutations in non-muscle γ-actin at the DFNA20/26 locus cause autosomal dominant nonsyndromic hearing loss. The molecular basis for the hearing loss is unknown. We have engineered each γ-actin mutation into yeast actin to investigate the effects of these mutations on actin function in vivo and in vitro. Cells expressing each of the mutant actins as the sole actin in the cell were viable. Four of the six mutant strains exhibited significant growth deficiencies in complete medium and an inability to grow on glycerol as the sole carbon source, implying a mitochondrial defect(s). These four strains exhibited abnormal mitochondrial morphology, although the mtDNA was retained. All of the mutant cells exhibited an abnormally high percentage of fragmented/non-polarized actin cables or randomly distributed actin patches. Five of the six mutants displayed strain-specific vacuole morphological abnormalities. Two of the purified mutant actins exhibited decreased thermal stability and increased rates of nucleotide exchange, indicative of increased protein flexibility. V370A actin alone polymerized abnormally. It aggregated in low ionic strength buffer and polymerized faster than wild-type actin, probably in part because of enhanced nucleation. Mixtures of wild-type and V370A actins displayed kinetic properties in proportion to the mole fraction of each actin in the mixture. No dominant effect of the mutant actin was observed. Our results suggest that a major factor in the deafness caused by these mutations is an altered ability of the actin filaments to be properly regulated by actin-binding proteins rather than an inability to polymerize.
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U2 - 10.1074/jbc.M601514200
DO - 10.1074/jbc.M601514200
M3 - Article
C2 - 16690605
AN - SCOPUS:33745989872
SN - 0021-9258
VL - 281
SP - 20129
EP - 20139
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 29
ER -