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Scaffold protein Receptor for Activated C Kinase 1 (RACK1) is a negative regulator of plant stress hormone - abscisic acid (ABA) mediated pathways. RACK1 has been reported to regulate global miRNA biogenesis pathway in C. elegans, humans, and in Arabidopsis. RACK1 regulates different steps of miRNA biogenesis and stability in response to different stimuli in plants. miR393s is implicated in salt stress response pathway through an antagonistic response between the stress hormone ABA-mediated salt stress and growth hormone auxin. Specifically, the known auxin receptor clade transcripts TIR1/AFB2 are the target for the miR393s. By down-regulating the auxin signaling pathways, the miR393s inhibit the regulation of salt tolerance by auxin. Here we show that genetic loss of RACK1A- the predominant member of the three genes family of RACK1 in Arabidopsis, results in the inhibition of miR393 level causing the same salt sensitivities as the individual mir393a or mir393b or the double mutant mir393ab phenotypes. We propose that down-regulation of auxin signaling through RACK1A induced miR393 biogenesis potentially regulates the Arabidopsis acclimation to salinity. Our findings fill up a molecular gap in our understanding of the role of miR393 mediated ABA and auxin-regulated salt stress responses.

After effectively eliminating the nonspecific cross-immunoreactivity with the affinity columns of anti-IgG agarose and IgG agarose, the potent immunoreactivities of p11 and calcyclin in wheat germ, lobster tail muscle, and three strains of baker's yeast were analyzed by Western blotting using mouse anti-p11 and rabbit anti-calcyclin. The occurrence of multiple bands may be due to either autolyses and/or the interactions between the p11 (or calcyclin) and other endogenous biological molecules. The results suggest not only a ubiquitous distribution and a universal Ca(2+)-mediating regulatory role of p11 and calcyclin in eukaryotes, but also an evolutionary conservation of these (S-100)-related proteins.

Vertebrate m-calpain, calpastatin, constitutive nitric oxide synthase, myelin basic protein, and dynamin I are substrates of protein kinase C (PKC). The presence/absence of similar/related protein in nonvertebrate was investigated by immunological methods, including (1) affinity chromatography on agarose-secondary antibodies and agarose IgG for removal of nonspecific immunoreactivities from crude extracts; (2) omitting beta-mercaptoethanol treatment and boiling prior to SDS-PAGE to increase the immunoreactivity; (3) immunoreactivity comparisons of nonspecific IgG as controls with specific anti-(vertebrate PKC-substrates/related proteins) in Western blots. It was found that (a) m-calpain and dynamin I were absent in baker's yeast, wheat germ and lobster tail muscle, (b) m-calpain, nitric oxide synthase, myelin basic protein and dynamin II were present in all three samples, and (c) calpastatin was present in baker's yeast and lobster tail muscle. The presence and absence of these proteins suggest evolutionary conservation and divergence, respectively, of these PKC substrates.

Varied immunoreactive bands of protein kinase C delta (PKC delta), PKC eta, and PKC zeta were detected in crude extracts of wheat germ, lobster tail meat, and three strains of baker's yeast by analysis of Western blots. Protease-deficient and Fleischmann's Active Dry yeasts exhibited immunoreactivity of PKC delta, whereas wheat germ and Fleischmann's RapidRise yeast displayed immunoreactivities of both PKC delta and PKC zeta. Lobster tail meat showed immunoreactivities of PKC eta and PKC zeta. These positive and negative immunoreactivities reflected evolutionary conservation and divergence, respectively, of these PKC isozymes in eukaryotes.

The immunoreactivity of S-100 proteins in three strains of yeast and wheat germ was observed by analysis of Western blotting with rabbit anti-bovine S-100. A high level of activity was exhibited in wheat germ, whereas a very low level was found in lobster tail meat. The occurrence of multiple bands may be due to the interactions between S-100A and S-100B and/or other molecules. The highly evolutionally conserved S-100 may play an important role in cellular signal transduction and cell growth in yeast and wheat germ.