dissection-local-ca2-signals-cultured-cells-membrane-targeted-ca2

Dissection of Local Ca<sup>2+</sup> Signals in Cultured Cells by Membrane-targeted Ca<sup>2+</sup> Indicators

Calcium ion (Ca2+) is a universal intracellular messenger molecule that drives multiple signaling pathways, leading to diverse biological outputs. The ...

RIKEN Center for Brain Science

Spinocerebellar ataxia type 29 (SCA29) is autosomal dominant congenital ataxia characterized by early-onset motor delay, hypotonia, and gait ataxia. Recently, heterozygous missense mutations in an intracellular Ca channel, inositol 1,4,5-trisphosphate (IP) receptor type 1 (IPR1), were identified as a cause of SCA29. However, the functional impacts of these mutations remain largely unknown. Here, we determined the molecular mechanisms by which pathological mutations affect IPR1 activity and Ca dynamics. Ca imaging using IPR-null HeLa cells generated by genome editing revealed that all SCA29 mutations identified within or near the IP-binding domain of IPR1 completely abolished channel activity. Among these mutations, R241K, T267M, T267R, R269G, R269W, S277I, K279E, A280D, and E497K impaired IP binding to IPR1, whereas the T579I and N587D mutations disrupted channel activity without affecting IP binding, suggesting that T579I and N587D compromise channel gating mechanisms. Carbonic anhydrase-related protein VIII (CA8) is an IPR1-regulating protein abundantly expressed in cerebellar Purkinje cells and is a causative gene of congenital ataxia. The SCA29 mutation V1538M within the CA8-binding site of IPR1 completely eliminated its interaction with CA8 and CA8-mediated IPR1 inhibition. Furthermore, pathological mutations in CA8 decreased CA8-mediated suppression of IPR1 by reducing protein stability and the interaction with IPR1. These results demonstrated the mechanisms by which pathological mutations cause IPR1 dysfunction, i.e., the disruption of IP binding, IP-mediated gating, and regulation via the IPR-modulatory protein. The resulting aberrant Ca homeostasis may contribute to the pathogenesis of cerebellar ataxia.

Matsumi Hirose

Laboratory for Developmental Neurobiology

RIKEN Center for Brain Science

Dissection of Local Ca²⁺ Signals in Cultured Cells by Membrane-targeted Ca²⁺ Indicators

Matsumi Hirose

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Laboratory for Developmental Neurobiology

RIKEN Center for Brain Science

Dissection of Local Ca2+ Signals in Cultured Cells by Membrane-targeted Ca2+ Indicators

Laboratory for Developmental Neurobiology

Dissection of Local Ca²⁺ Signals in Cultured Cells by Membrane-targeted Ca²⁺ Indicators