Zhong-Ling Zhuo1, Lu Cong2, Jun Zhang2, Xiao-Tao Zhao1. 1. Department of Clinical Laboratory, Peking University People's Hospital, Beijing, China. 2. Department of Neurology, Peking University People's Hospital, Beijing, China.
Abstract
BACKGROUND: We investigated whether a heterozygous mutation that we newly identified in HTRA1 (high-temperature requirement serine protease A1 gene) in a pedigree with autosomal dominant hereditary cerebral small vessel disease (SVD) reduces the function of HTRA1 and affects the transforming growth factor-β1 (TGF-β1)/Smad signaling. METHODS: Whole-exome sequence from the proband and her two sisters was examined using whole-exome enrichment and sequencing. Expression of HTRA1 and TGF-β1/Smad and HTRA1 activity were assayed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting analyses after transfecting wild-type and mutant HTRA1 genes into HEK293 cells. RESULTS: A new heterozygous mutation (c.614C>G:p.Ser205Cys) in HTRA1 was identified in the sequence encoding the trypsin-like serine protease domain. The mutation was predicted to be deleterious by in silico tools. Moreover, in vitro activity and protein analyses revealed a loss-of-function effect of the mutation: the proteolytic activity of mutant HTRA1 was decreased, and, notably, this was accompanied by an increase in TGF-β1/Smad protein levels. CONCLUSIONS: The heterozygous mutation HTRA1 S205C causing diminished protease activity is associated with-and could represent a cause of-autosomal dominant hereditary cerebral SVD. Our results also indicate a relationship between HTRA1 and TGF-β1/Smad signaling.
BACKGROUND: We investigated whether a heterozygous mutation that we newly identified in HTRA1 (high-temperature requirement serine protease A1 gene) in a pedigree with autosomal dominant hereditary cerebral small vessel disease (SVD) reduces the function of HTRA1 and affects the transforming growth factor-β1 (TGF-β1)/Smad signaling. METHODS: Whole-exome sequence from the proband and her two sisters was examined using whole-exome enrichment and sequencing. Expression of HTRA1 and TGF-β1/Smad and HTRA1 activity were assayed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting analyses after transfecting wild-type and mutant HTRA1 genes into HEK293 cells. RESULTS: A new heterozygous mutation (c.614C>G:p.Ser205Cys) in HTRA1 was identified in the sequence encoding the trypsin-like serine protease domain. The mutation was predicted to be deleterious by in silico tools. Moreover, in vitro activity and protein analyses revealed a loss-of-function effect of the mutation: the proteolytic activity of mutant HTRA1 was decreased, and, notably, this was accompanied by an increase in TGF-β1/Smad protein levels. CONCLUSIONS: The heterozygous mutation HTRA1S205C causing diminished protease activity is associated with-and could represent a cause of-autosomal dominant hereditary cerebral SVD. Our results also indicate a relationship between HTRA1 and TGF-β1/Smad signaling.