Cerebral autosomal-recessive arteriopathy with subcortical infarcts and leukoencephalopathy (CARASIL), an autosomal-recessive inherited cerebral small vessel disease (CSVD), involves severe leukoaraiosis, multiple lacunar infarcts, early-onset alopecia, and spondylosis deformans without hypertension. High-temperature requirement serine peptidase A1 (HTRA1) gene mutations cause CARASIL by decreasing HTRA1 protease activity. Although CARASIL is a recessive inherited disease, for some HTRA1 mutations, the parents of the probands with CARASIL present mild to severe white matter lesions. Very recently, heterozygous mutations in HTRA1 were identified in 11 families with CSVD; so far, most of the parents of CARASIL patients with HTRA1 missense mutations, who are assumed to be heterozygotes, did not show symptomatic CSVD. Moreover, heterozygotes carrying the nonsense mutations, in whom residual HTRA1 activity might be near 50% that of controls, did not show symptomatic CSVD. Therefore, it is unclear which mutants truly contribute to CSVD pathogenesis and how HTRA1 mutants cause CSVD in heterozygous individuals. Here we found four heterozygous missense mutations in the HTRA1 gene (p.G283E, p.P285L, p.R302Q, and p.T319I) in six patients from 113 unrelated index patients. An autopsied case with p.G283E showed arteriopathy in their cerebral small arteries. These mutant HTRA1s showed markedly decreased protease activities and inhibited wild-type HTRA1 activity, whereas two of three mutant HTRA1s reported in CARASIL did not inhibit wild-type HTRA1 activity. Wild-type HTRA1 forms trimers; however, G283E and T319I HTRA1, observed in manifesting heterozygotes, did not form trimers. P285L and R302Q HTRA1s formed trimers, but their mutations were located in domains that are important for trimer-associated HTRA1 activation; in contrast, mutant HTRA1s observed in CARASIL also formed trimers but had mutations outside the domains important for trimer-associated HTRA1 activation.
The mutant HTRA1s observed in manifesting heterozygotes might result in an impaired HTRA1 activation cascade of HTRA1 or be unable to form stable trimers. These data have demonstrated that the mutant HTRA1s associated with manifesting heterozygotes have dominant-negative effects on wild-type HTRA1 protease activity, indicating that the HTRA1 protease activities of manifesting heterozygotes might be less than those of heterozygotes carrying the nonsense mutations that do not develop symptomatic CSVD.

研究助成：Research funds : a grant-in-aid for Scientific Research on Innovative Areas (Brain Protein Aging and Dementia Control) (26117006) from MEXT; a gr