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  1. Fulltext Contribution of Coiled-Coil Assembly to Ca2+/Calmodulin-Dependent Inactivation of TRPC6 Channel and its Impacts on FSGS-Associated Phenotypes
    Polat OK, Uno M, Maruyama T, Tran HN, Imamura K, Wong CF, et al.
    J Am Soc Nephrol, 2019 09;30(9):1587-1603.
    PMID: 31266820 DOI: 10.1681/ASN.2018070756
    BACKGROUND: TRPC6 is a nonselective cation channel, and mutations of this gene are associated with FSGS. These mutations are associated with TRPC6 current amplitude amplification and/or delay of the channel inactivation (gain-of-function phenotype). However, the mechanism of the gain-of-function in TRPC6 activity has not yet been clearly solved.

    METHODS: We performed electrophysiologic, biochemical, and biophysical experiments to elucidate the molecular mechanism underlying calmodulin (CaM)-mediated Ca2+-dependent inactivation (CDI) of TRPC6. To address the pathophysiologic contribution of CDI, we assessed the actin filament organization in cultured mouse podocytes.

    RESULTS: Both lobes of CaM helped induce CDI. Moreover, CaM binding to the TRPC6 CaM-binding domain (CBD) was Ca2+-dependent and exhibited a 1:2 (CaM/CBD) stoichiometry. The TRPC6 coiled-coil assembly, which brought two CBDs into adequate proximity, was essential for CDI. Deletion of the coiled-coil slowed CDI of TRPC6, indicating that the coiled-coil assembly configures both lobes of CaM binding on two CBDs to induce normal CDI. The FSGS-associated TRPC6 mutations within the coiled-coil severely delayed CDI and often increased TRPC6 current amplitudes. In cultured mouse podocytes, FSGS-associated channels and CaM mutations led to sustained Ca2+ elevations and a disorganized cytoskeleton.

    CONCLUSIONS: The gain-of-function mechanism found in FSGS-causing mutations in TRPC6 can be explained by impairments of the CDI, caused by disruptions of TRPC's coiled-coil assembly which is essential for CaM binding. The resulting excess Ca2+ may contribute to structural damage in the podocytes.

    Matched MeSH terms: Calmodulin/genetics
  2. Molecular epidemiology of Sporothrix schenkii isolates in Malaysia
    Kamal Azam NK, Selvarajah GT, Santhanam J, Abdul Razak MF, Ginsapu SJ, James JE, et al.
    Med Mycol, 2020 Jul 01;58(5):617-625.
    PMID: 31642485 DOI: 10.1093/mmy/myz106
    Sporothrix schenkii is a dimorphic fungus that causes infections in both humans and animals. We report on 25 S. schenkii isolates collected in 2017 from humans and cats clinically diagnosed with sporotrichosis, in Malaysia. These isolates were phenotypically identified as S. schenkii sensu lato and further defined as S. schenckii sensu stricto based on partial calmodulin gene sequence. Isolates from both humans and cats were genotypically identical but displayed phenotypic variation. Phylogenetic analyses based on partial calmodulin sequence showed that the Malaysian isolates clustered with global S. schenkii sensu stricto strains, in particular, of the AFLP type E. This analysis also revealed that partial calmodulin sequence alone was sufficient for classifying global S. schenckii sensu stricto strains into their respective AFLP types, from A to E. The genetically conserved S. schenkii sensu stricto species isolated from humans and cats is suggestive of a clonal strain present in Malaysia. To the best of our knowledge, this is the first report on molecular identification of Sporothrix schenkii strains from human infections in Malaysia. Further studies are required in order to elucidate the clonal nature of Malaysian S. schenkii isolates. Our findings indicate the presence of a predominant S. schenkii genotype in the environment, causing infections in both cats and humans in Malaysia.
    Matched MeSH terms: Calmodulin/genetics
  3. Molecular typing of Sporothrix schenckii isolates from cats in Malaysia
    Kano R, Okubo M, Siew HH, Kamata H, Hasegawa A
    Mycoses, 2015 Apr;58(4):220-4.
    PMID: 25727965 DOI: 10.1111/myc.12302
    Epidemiological data on the aetiologic agents of feline sporotrichosis in Malaysia have not been reported, though human sporotrichosis in Malaysia is reported to be transmitted primarily via cat scratch. To the best of our knowledge, the present report is the first study of the molecular epidemiology of Sporothrix schenckii isolates from cats with sporotrichosis in Malaysia. In the present work, we characterised 18 clinical isolates from cats in Malaysia based on molecular properties, including sequence analyses of the calmodulin gene and the rDNA ITS region and selective PCR of mating type (MAT) loci. In this study, isolates from feline sporotrichosis were identified as a S. schenckii sensu stricto by sequence analyses of the calmodulin gene and the internal transcribed spacer (ITS) region. Notably, phylogenetic analysis of the ITS confirmed assignment to clinical clade D (and not C) of S. schenckii sensu stricto. Therefore, clinical clade D of S. schenckii sensu stricto appeared to be the prevailing source of feline sporotrichosis in Malaysia. The ratio of MAT1-1-1:MAT1-2-1 in these Malaysian isolates was found to be 1 : 0. This result suggested that a clonal strain of S. schenckii is the prevailing causative agent of feline sporotrichosis in Malaysia.
    Matched MeSH terms: Calmodulin/genetics
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