Distinct Calcium Signaling Pathways Regulate Calmodulin Gene Expression in Tobacco

Abstract

Cold shock and wind stimuli initiate Ca²⁺ transients in transgenic tobacco (Nicotiana plumbaginifolia) seedlings (named MAQ 2.4) containing cytoplasmic aequorin. To investigate whether these stimuli initiate Ca²⁺ pathways that are spatially distinct, stress-induced nuclear and cytoplasmic Ca²⁺ transients and the expression of a stress-induced calmodulin gene were compared. Tobacco seedlings were transformed with a construct that encodes a fusion protein between nucleoplasmin (a major oocyte nuclear protein) and aequorin. Immunocytochemical evidence indicated targeting of the fusion protein to the nucleus in these plants, which were named MAQ 7.11. Comparison between MAQ 7.11 and MAQ 2.4 seedlings confirmed that wind stimuli and cold shock invoke separate Ca²⁺ signaling pathways. Partial cDNAs encoding two tobacco calmodulin genes, NpCaM-1 andNpCaM-2, were identified and shown to have distinct nucleotide sequences that encode identical polypeptides. Expression ofNpCaM-1, but not NpCaM-2, responded to wind and cold shock stimulation. Comparison of the Ca²⁺dynamics with NpCaM-1 expression after stimulation suggested that wind-induced NpCaM-1 expression is regulated by a Ca²⁺ signaling pathway operational predominantly in the nucleus. In contrast, expression ofNpCaM-1 in response to cold shock is regulated by a pathway operational predominantly in the cytoplasm.