tubule motor protein, KCBP, containing a CaM-binding domain at its C-terminus and other unique do-
mains in the N-terminus has been identified from Arabidopsis[218,236], tobacco [242], and potato
[237,268–272]. Furthermore, the ArabidopsisCaM isoforms show differential affinity for this motor pro-
tein [218]. Disruption of KCBP revealed its involvement in trichome cell shape morphology in Ara-
bidopsis[273,274]. Immunolocalization and microinjection studies indicate a role for KCBP in cell divi-
sion [240,241].
Using differential screening of cDNA libraries prepared from NaCl-treated and-untreated tomato
seedlings, a cDNA was isolated for glyoxylase I [245] and its transcripts are inducible by either NaCl,
mannitol, or abscisic acid treatments in tomato [245]. Glyoxalase I, which catalyzes the conversion of
toxic methylglyoxal to nontoxic metabolite in plants, has been purified using CaM-Sepharose affinity col-
umn chromatography from Brassica juncea[275]. Its activity is stimulated by Ca^2 alone (2.6-fold) or
Ca^2 /CaM (5 M/145 nM) (7-fold). Further, the concentration of CaM required to stimulate the glyox-
alase activity to the same extent in the presence of Mg^2 (29 nM) and Ca^2 (145 nM) is varied, suggest-
ing that CaM has a cumulative effect on the metal-dependent activation of glyoxalase [275]. Using anti-
bodies raised against purified B. junceaglyoxalase I as a probe, a cDNA sequence has been isolated from
the cDNA expression library constructed for mRNA isolated from mannitol-treated B. juncea[244]. The
full-length glyoxalase cDNA (BjGlyI, Figure 3) is 784 bp in length and consists of an 558-bp ORF [185
amino acids (aa)]. Although Ca^2 /CaM stimulate its activity, the CaM-binding domain is not mapped.
The binding sites of two cofactors (Zn^2 and glutathione) along with the putative serine/threonine phos-
phorylation sites are shown to be conserved in glyoxalase I [244]. Further, its enzyme activity is stimu-
lated by stress such as polyethylene glycol (PEG), light, and phytohormones in B. juncea[276].
Harrington and his coworkers [246] have successfully isolated several putative cDNA clones en-
coding CaMBPs by screening an expression library constructed from cell cultures of heat-shocked to-
bacco with [^35 S]calmodulin. Of these, one cDNA sequence encodes a 499-amino-acid long protein,
NtCBP48 (Figure 3). The sequence analysis predicts that it contains a centrally located transmembrane
domain and nuclear localization sequence motif [246]. Using bacterially expressed deletion mutants, pu-
rification through CaM-affinity chromatography, and their interaction with CaM on gel mobility shift as-
say, these authors have mapped the CaM-binding domain of CBP48 to a region of 40 amino acids in the
C-terminus (Figure 3). The corresponding gene transcripts are rapidly inducible by heat shock (38°C)
reaching maximum in 1.5 hr after treatment, indicating a role for NtCBP48 in heat stress [246].
C. Other Calcium-Binding Proteins Involved in Stress
A new family of Ca^2 -binding proteins, called calcineurin B–like (CBL) proteins, has been identified
fromArabidopsis[63,204,277]. They are similar to the regulatory B subunit of calcineurin and the neu-
ronal Ca^2 sensor (NCS). Studies with Arabidopsisrevealed the presence of at least six genes encoding
highly similar but functionally distinct AtCBL proteins [204]. Although CaM and CBLs belong to two
different groups, CBLs also contain four EF hand motifs and their activity depends on Ca^2 binding (Fig-
ure 2). Drought, cold, and wound stress signals elevate AtCBL1gene transcripts, whereas AtCBL2and
AtCBL3are constitutively expressed [204]. The AtCBL4(also called SOS3) is involved in the salt stress
tolerance mechanism [63]. Further, CBL-interacting protein kinases (CIPK1 to 4) have also been identi-
fied using a yeast two-hybrid screen [277]. These protein kinases belong to the serine/threonine class of
kinases and show high homology to SNF1 and AMPK from yeast and mammalian systems, respectively.
These CIPKs interact with CBLs, but not with CaMs, in a Ca^2 -dependent manner [277]. Several iso-
forms of AtCBLs and CIPKs were also identified. However, CBL-CIPK isoform specificity is not deter-
mined and future studies in this direction should open up new avenues of research.
Using differential screening of a rice cDNA library constructed from ABA-treated rice seedlings, a
cDNA sequence encoding a 27.4-kDa protein, EFA27, has been isolated [205]. It contains a single con-
served EF hand motif, a characteristic Ca^2 -binding domain (Figure 2). Sequence analysis and a Ca^2 -
binding assay with E. coli–expressed and–purified protein indicate that it is a new member of a Ca^2 -
binding protein family that is induced by ABA and osmotic stress [205]. The EFA27gene transcripts are
inducible in response to salt and dehydration stress and to an ABA signal. Sequence homology searches
indicate that there are several EFA27gene homologues in Arabidopsis[205], suggesting the existence of
similar proteins in phylogenetically distant species. Further, EF hand motif–containing protein phos-
phatases were also identified in the ABA signaling pathway [278]. Another Ca^2 -binding protein, AtCP1,
CALCIUM IN STRESS SIGNAL TRANSDUCTION 711