plant growth and development. This phenotype is due to anatomical abnormalities in cell elongation in
stem cortex and parenchyma tissues. This phenotype was shown to correlate with accumulation of ex-
tremely high levels of GABA and low levels of Glu [232]. This is a very important observation in light
of the plants’ ability to produce high levels of GABA in response to a variety of stresses [264,351].
Fromm and his colleagues [258] have isolated a cDNA sequence, NtCBP4, encoding a protein with
a CaM-binding property from tobacco. Sequence comparison indicates its close resemblance to nonse-
lective membrane-bound cation channels from animals, its plant homologues were isolated from Ara-
bidopsis[352] and barley [234], and it has been shown that it is located on plasma membrane [234,258].
Constitutive expression of NtCBP4cDNA under the 35S promoter in sense and antisense orientation in
tobacco exhibited a normal phenotype under normal growth conditions. However, when subjected to dif-
ferent metal toxicities, NtCBP4 antisense and GAD-expressing transgenics along with the wild-type
plants showed symptoms associated with Ni^2 toxicity such as retarded root and shoot growth and re-
duced chlorophyll content (chlorotic) although all contained the same amount of NtCBP4 protein. In con-
trast, the NtCBP4sense transgenic plants grew and developed normally even at 200 M NiCl 2. However,
sense plants showed hypersensitivity to Pb^2 [258]. This is the first report that a protein (NtCBP4) show-
ing CaM-binding capacity is involved in metal tolerance. Further, the CBD is speculated to be located at
the C-terminus near the end of the cyclic nucleotide binding site (Figure 3).
Another example of involvement of Ca^2 /CaM regulation in the plant disease resistance response
comes from plants expressing divergent CaMs [220]. Constitutive expression of divergent CaM isoforms
SCaM4 and -5 under the 35S promoter in tobacco has yielded the important biological functional signif-
icance of the most divergent CaM isoforms identified so far. Conserved CaMs (SCaM1, -2, and -3) are
not inducible by stress, whereas divergent CaM isoforms are normally expressed at a low level and are
highly inducible in response to Ca^2 -mediated pathogen attack (Fusarium solani,Phytophthora parasit-
icavarnicotianae) or elicitor contact (derived from P. parasitica). Transgenic plants expressing high lev-
els of SCaM4 and -5 showed constitutive expression of SA-related gene expression independent of SA
throughout their life cycle and showed enhanced disease resistance to a wide spectrum of pathogens (Phy-
tophthora parasiticavarnicotiana,Pseudomonas syringaepv.tabaci, and a virulent viral pathogen,
TMV) [220]. These studies revealed the existence of CaM isoform specificity in their target activation in
Ca^2 -mediated signal transduction processes in plants. Furthermore, studies revealed that the CaM iso-
forms differ in their affinity for the same target protein [106,167,174,192,193,201,218,316], highlighting
the significance of the existence of multiple CaM isoforms in mediating specific Ca^2 -mediated signal
transduction pathways. However, the CaM target proteins involved in various processes remain to be
characterized.
In order to understand the biological significance of a small CaM-binding protein, BjGLY 1, Veena
et al. [244] raised transgenic tobacco with the BjGly Igene under the CaMV35S promoter. The transgenic
plants showed a normal phenotype as antisense BjGly Itransgenic and wild-type plants. However, trans-
genic plants overexpressing the BjGly 1gene showed elevated levels of BjGly Igene transcripts when
subjected to NaCl (50 to 800 mM), mannitol (100 to 800 mM), or ZnCl 2 [244]. Further, leaf disks of trans-
genic plants were assayed for their tolerance to abiotic stresses (NaCl, mannitol, methylglyoxal). The re-
sults revealed that the sense transgenic leaf disks showed tolerance with an intact chlorophyll content and
without any deterioration from the normal phenotype. In contrast, the control antisense and wild-type leaf
disks showed a bleaching effect (loss of chlorophyll) and did not survive these stresses. These results sug-
gest that BjGly I plays a role in conferring tolerance to salt, methyl glyoxal, and water stresses (abiotic)
in plants.
VI. CONCLUSIONS
The reports described in this chapter clearly indicate an important messenger role for Ca^2 in transduc-
ing a variety of stress signals. Stress-induced rapid and transient changes in cytosolic Ca^2 have been doc-
umented in several cases. At least two classes of proteins (CaM and CDPK) are found in all the plants that
seem to play a broad role in mediating Ca^2 action. Several other Ca^2 -sensing proteins (calcineurin B-
like, channels, antiporters) that mediate Ca^2 stress tolerance or regulate Ca^2 changes in the cytoplasm
have been functionally characterized. In addition, several proteins that are downstream targets of
Ca^2 /CaM have been detected, and in some cases their involvement in stress-related processes has been
720 REDDY AND REDDY