The Grapevine Calmodulin-Like Protein Gene CML21 Is Regulated by Alternative Splicing and Involved in Abiotic Stress Response

Abstract

Calmodulin-like proteins (CMLs) represent a large family of plant calcium sensor proteins involved in the regulation of plant responses to environmental cues and developmental processes. In the present work, we identified four alternatively spliced mRNA forms of the grapevine CML21 gene that encoded proteins with distinct N-terminal regions. We studied the transcript abundance of CML21v1, CML21v2, CML21v3, and CML21v4 in wild-growing grapevine Vitis amurensis Rupr. in response to desiccation, heat, cold, high salinity, and high mannitol stress using quantitative real-time RT-PCR. The levels of all four splice variants of VaCML21 were highly induced in response to cold stress. In addition, VaCML21v1 and VaCML21v2 forms were highly modulated by all other abiotic stress treatments. Constitutive expression of VaCML21v2 and VaCML21v4 improved biomass accumulation of V. amurensis callus cell cultures under prolonged low temperature stress. Heterologous expression of the grapevine CML21v2 and VaCML21v4 splice variants in Arabidopsis improved survival rates of the transgenic plants after freezing. The VaCML21v2 overexpression enhanced activation of the cold stress-responsive marker genes AtDREB1A and AtDREB2A, while VaCML21v4 overexpression—AtCOR47, AtRD29A, AtRD29B, and AtKIN1 genes after freezing stress in the transgenic Arabidopsis. The results indicate that the grapevine CML21 gene acts as a positive regulator in the plant response to cold stress. The detected variety of CML21 transcripts and their distinct transcriptional responses suggested that this expansion of mRNA variants could contribute to the diversity of grapevine adaptive reactions.

Transcriptomics Reveals the ERF2-bHLH2-CML5 Module Responses to H2S and ROS in Postharvest Calcium Deficiency Apples

“…There are numerous studies reporting that CMLs are involved in regulating a variety of abiotic stress processes. For example, SlCML37 enhances cold tolerance in tomato fruit [ 39 ]; CmCML13 improves drought resistance in Arabidopsis [ 40 ]; CML21 is involved in abiotic stress response in grapevine [ 41 ]. Thus, it can be hypothesized that CML5 may be involved in regulating postharvest storage quality and antioxidant-related processes in calcium-deficient apple peels.…”

Physiological Conditions and dsRNA Application Approaches for Exogenously induced RNA Interference in Arabidopsis thaliana

“…The isolation of total RNA was performed using the cetyltrimethylammonium bromide (CTAB)-based protocol [ 45 ] and complementary DNAs were synthesized as described [ 46 ].…”

External dsRNA Downregulates Anthocyanin Biosynthesis-Related Genes and Affects Anthocyanin Accumulation in Arabidopsis thaliana

“…For RNA isolations, a typical adult leaf of A. thaliana was collected from the same individual plant before treatment, two days and seven days post-treatment for each type of treatment in an independent experiment. Total RNA was isolated using the cetyltrimethylammonium bromide (CTAB)-based protocol [56] and complementary DNAs were synthesized as described [57].…”