CAMTA3/SR1 (herein CAMTA3) functions as a repressor of gene expression of defense responses (). More recently, two research groups independently identified mutations suppressing defense responses and systemic acquired resistance (SAR), conferred by the same amino acid substitution (A855V) in the IQ-I domain of CAMTA3 (). This mutation conferred enhanced susceptibility to pathogens suggesting that the binding of CAMTA3 to its targets is tighter, and thus enhances repression of gene expression. In vitro tests showed that a truncated protein containing the CAMTA3 IQ domains with or without the mutation bound CaM in a Ca-dependent manner (). The mutation A855V could also possibly enhance the binding of CaM to CAMTA3 through the C-terminal region where both IQ and CaMBD are located close together, and subsequently may enhance Ca/CaM-mediated repression of target gene expression, a scenario suggested by, corresponding to Figure 1 C. Interestingly, structural analysis of CaM binding to the IQ1 domain of myosin () revealed that in the absence of Cathe C-terminal lobe of CaM binds to the IQ1 motif but the N-terminal lobe of CaM is unbound. Upon Cabinding to CaM and the concomitant conformational change, the N-terminal lobe of CaM may bind to amino acids adjacent to the IQ domain, the precise site being dependent on the composition of the amino acids within and adjacent to the IQ domain. Moreover, IQ domains are often found adjacent to canonical Ca-dependent CaM binding domains, as in the case of plant CAMTA3 and CNGCs ().

Striped rectangles denote gene promoters; red spheres denote Caions. The number of red spheres denotes whether the calcium concentration is expected to be low (one red sphere) or high (4 red spheres).Represent hypothetical situations. In, ubiquitin-mediated degradation was reported by, whereas Ca/CaM-dependent removal of the TF from the promoter is hypothetical.

In calmodulin-IQ domain complexes, the Ca 2+ -free and Ca 2+ -bound forms of the calmodulin C-lobe direct the N-lobe to different binding sites.

Du et al., 2009 Du L.

Ali G.S.

Simons K.A.

Hou J.

Yang T.

Reddy A.S.

Poovaiah B.W. Ca2+/calmodulin regulates salicylic-acid-mediated plant immunity.

Truman et al. (2013) Truman W.

Sreekanta S.

Lu Y.

Bethke G.

Tsuda K.

Katagiri F.

Glazebrook J. The calmodulin-binding protein 60 family includes both negative and positive regulators of plant immunity.

Miller et al., 2013 Miller J.B.

Pratap A.

Miyahara A.

Zhou L.

Bornemann S.

Morris R.J.

Oldroyd G.E. Calcium/calmodulin-dependent protein kinase is negatively and positively regulated by calcium, providing a mechanism for decoding calcium responses during symbiosis signaling.

Black and Persechini, 2011 Black D.J.

Persechini A. In calmodulin-IQ domain complexes, the Ca2+-free and Ca2+-bound forms of the calmodulin C-lobe direct the N-lobe to different binding sites.

Nie et al., 2012 Nie H.

Zhao C.

Wu G.

Wu Y.

Chen Y.

Tang D. SR1, a calmodulin-binding transcription factor, modulates plant defense and ethylene-induced senescence by directly regulating NDR1 and EIN3.

Du et al., 2009 Du L.

Ali G.S.

Simons K.A.

Hou J.

Yang T.

Reddy A.S.

Poovaiah B.W. Ca2+/calmodulin regulates salicylic-acid-mediated plant immunity.

Truman et al., 2013 Truman W.

Sreekanta S.

Lu Y.

Bethke G.

Tsuda K.

Katagiri F.

Glazebrook J. The calmodulin-binding protein 60 family includes both negative and positive regulators of plant immunity.

Zhang et al., 2014 Zhang L.

Du L.

Shen C.

Yang Y.

Poovaiah B.W. Regulation of plant immunity through ubiquitin-mediated modulation of Ca2+-calmodulin-AtSR1/CAMTA3 signaling.

Furthermore, whereas binding of a truncated CAMTA3 (CG-1 domain fragment) to the EDS1 promoter in vitro is Ca/CaM independent (), mutations in the CaM binding domain of CAMTA3, which abolish CaM binding in vitro (e.g., K907E), are unable to repress EDS1 expression in vivo. The authors interpreted these results as evidence that CaM binding to CAMTA3 is required for its role as a transcriptional repressor. However, no direct evidence for the role of Cain this process has been provided, and the possibility that the mutations affected the overall protein conformation causing reduced binding of CAMTA3 to the EDS1 promoter irrespective of CaM binding cannot be ruled out. Similarly,suggested that CaM binding to the CBP60a is required for its role in repression of its target gene (SID2) in the immune response, and questioned whether this could be dependent on Cain the absence of pathogen attack. The authors suggested that “there must be enough Caand CaM present in the absence of pathogens to allow function of the CBP60a, CAMTA and CBNAC repressors”. While this explanation cannot be ruled out, the fact that nuclear Calevels are expected to be low prior to pathogen attack raises questions about the occurrence of Ca-dependent transcriptional repression mechanisms, both in the case of CBP60a and CAMTA3. Alternatively, instead of considering Ca-dependent versus Ca-independent CaM interactions with TFs, the in vivo CaM binding mechanism may be more complex by responding quantitatively to the local changes in sub-nuclear Calevels, or to the nature of the Casignal by conferring different Ca-dependent CaM–TF conformational changes (), perhaps similar to the case of CaM–myosin interactions described above (). Such subtle in vivo mechanisms would be difficult to observe in vitro with truncated proteins or with partial protein complexes, particularly when in vitro studies of Ca/CaM interactions are typically performed either in the presence of 1 mM Caor 5 mM EGTA (e.g.,). Therefore, recently suggested models of Ca-dependent transcriptional repression by CAMTA3 and CBP60a () need to be considered with caution. Figure 1 depicts possible mechanisms of Caand/or CaM dependent and independent transcription repression mechanisms.