Disulfide Dimerization of Neuronal Calcium Sensor-1: Implications for Zinc and Redox Signaling

Viktoriia E Baksheeva (1) , Alexey V Baldin (1) , Arthur O Zalevsky (2) , Aliya A Nazipova (2) , Alexey S Kazakov (2) , Vasiliy I Vladimirov (2) , Neonila V Gorokhovets (3) , François Devred (4) , Pavel P Philippov (1) , Alexandr V Bazhin (5) , Andrey V Golovin (2) , Andrey A Zamyatnin (1) , Dmitry V Zinchenko (2) , Philipp O Tsvetkov (4) , Sergei E Permyakov (2) , Evgeni Yu Zernii (1)

1 MSU - Lomonosov Moscow State University
2 RAS - the Russian Academy of Sciences [Moscow, Russia]
3 Sechenov First Moscow State Medical University
4 INP - Institut de neurophysiopathologie
5 DKTK - German Cancer Consortium [Heidelberg]

Neuronal calcium sensor-1 (NCS-1) is a four-EF-hand ubiquitous signaling protein modulating neuronal function and survival, which participates in neurodegeneration and carcinogenesis. NCS-1 recognizes specific sites on cellular membranes and regulates numerous targets, including G-protein coupled receptors and their kinases (GRKs). Here, with the use of cellular models and various biophysical and computational techniques, we demonstrate that NCS-1 is a redox-sensitive protein, which responds to oxidizing conditions by the formation of disulfide dimer (dNCS-1), involving its single, highly conservative cysteine C38. The dimer content is unaffected by the elevation of intracellular calcium levels but increases to 10–30% at high free zinc concentrations (characteristic of oxidative stress), which is accompanied by accumulation of the protein in punctual clusters in the perinuclear area. The formation of dNCS-1 represents a specific Zn2+-promoted process, requiring proper folding of the protein and occurring at redox potential values approaching apoptotic levels. The dimer binds Ca2+ only in one EF-hand per monomer, thereby representing a unique state, with decreased α-helicity and thermal stability, increased surface hydrophobicity, and markedly improved inhibitory activity against GRK1 due to 20-fold higher affinity towards the enzyme. Furthermore, dNCS-1 can coordinate zinc and, according to molecular modeling, has an asymmetrical structure and increased conformational flexibility of the subunits, which may underlie their enhanced target-binding properties. In HEK293 cells, dNCS-1 can be reduced by the thioredoxin system, otherwise accumulating as protein aggregates, which are degraded by the proteasome. Interestingly, NCS-1 silencing diminishes the susceptibility of Y79 cancer cells to oxidative stress-induced apoptosis, suggesting that NCS-1 may mediate redox-regulated pathways governing cell death/survival in response to oxidative conditions.

Mots clés: EF-hand; NCS family; neuronal calcium sensor-1; disulfide dimerization; GRK1; zinc; protein aggregation; apoptosis; neurodegeneration; cancer

Domaines: Biophysique; Neurobiologie; Cancer

Viktoriia E Baksheeva, Alexey V Baldin, Arthur O Zalevsky, Aliya A Nazipova, Alexey S Kazakov, et al.. Disulfide Dimerization of Neuronal Calcium Sensor-1: Implications for Zinc and Redox Signaling. International Journal of Molecular Sciences, 2021, Molecular Basis of Sensory Transduction in Health and Disease, ⟨10.3390/ijms222212602⟩. ⟨hal-03441623⟩