Genome-wide identification and biochemical characterization of calcineurin B-like calcium sensor proteins in Chlamydomonas reinhardtii

Abstract

Calcium (Ca2+) signaling is involved in the regulation of diverse biological functions through association with several proteins that enable them to respond to abiotic and biotic stresses. Though Ca2+-dependent signaling has been implicated in the regulation of several physiological processes in Chlamydomonas reinhardtii, Ca2+ sensor proteins are not characterized completely. C. reinhardtii has diverged from land plants lineage, but shares many common genes with animals, particularly those encoding proteins of the eukaryotic flagellum (or cilium) along with the basal body. Calcineurin, a Ca2+/calmodulin-dependent protein phosphatase, is an important effector of Ca2+ signaling in animals, while calcineurin B-like proteins (CBLs) play an important role in Ca2+ sensing and signaling in plants. The present study led to the identification of 13 novel CBL-like Ca2+ sensors in C. reinhardtii genome. One of the archetypical genes of the newly identified candidate, CrCBL-like1 was characterized. The ability of CrCBL-like1 protein to sense as well as bind Ca2+ were validated using two-step Ca2+-binding kinetics. The CrCBL-like1 protein localized around the plasma membrane, basal bodies and in flagella, and interacted with voltage-gated Ca2+ channel protein present abundantly in the flagella, indicating its involvement in the regulation of the Ca2+ concentration for flagellar movement. The CrCBL-like1 transcript and protein expression were also found to respond to abiotic stresses, suggesting its involvement in diverse physiological processes. Thus, the present study identifies novel Ca2+ sensors and sheds light on key players involved in Ca2+signaling in C. reinhardtii, which could further be extrapolated to understand the evolution of Ca2+ mediated signaling in other eukaryotes.

Genome-Wide Analysis of the Apple CBL Family Reveals That Mdcbl10.1 Functions Positively in Modulating Apple Salt Tolerance

“…[2][3][4][5]. Under stress conditions, Ca 2+ signals induced by the stimulus are first perceived, decoded, and transmitted by Ca 2+ sensors [6].…”

“…Ca 2+ can bind to the elongation factor (EF) hand domains of the CBL proteins, which changes their phosphorylation status. The change in the phosphorylation status of Ca 2+ sensors activates several protein kinases, which sometimes lead to a protein phosphorylation cascade [6,20,21].…”

Chlamydomonas reinhardtii cellular compartments and their contribution to intracellular calcium signalling

“…Interestingly, PKD2 was recently reported to be attached to mastigonemes, extracellular polymers of the glycoprotein mastigoneme-like protein 1 (MST1), and the axoneme, suggesting its mechanosensory role in motile cilia ( Liu et al , 2020 ). In addition, three Ca 2+ sensor kinases have been identified in the flagella proteome, namely CDPK1, CDPK3, and CDPK11, having a crucial role in the flagella length assembly and length control, whereas a Ca 2+ -binding CBL-like1 protein was recently also locali//////////zed in flagella, interacting with VGCCs ( Table 1 ) ( Pazour et al , 2005 ; Liang and Pan, 2013 ; Kumar et al , 2020 ). Such confined localization of these molecular players further supports the experimental evidence that a C. reinhardtii single flagellum can act as an independent Ca 2+ signalling entity from the cell body.…”

“… Li et al , 2019 ). Recently, 13 novel CBL-like proteins have also been identified through a genome-wide analysis in C. reinhardtii , displaying characteristic EF-hand Ca 2+ -binding domains like those present in Arabidopsis CBLs ( Kumar et al , 2020 ).…”