SummaryFlooding of maize seedlings causes O 2 deprivation that leads to a global reduction in protein synthesis and selective translation of cytoplasmic mRNAs. Since selective translation in animal cells can involve the capbinding protein eIF4E, we characterized the distinct mRNA cap-binding proteins eIF4E and eIFiso4E of maize. These proteins have 45% deduced amino acid sequence identity and are highly conserved at residues of eIF4E that function in intermolecular interactions in animals. Maize eIF4E is a phosphoprotein. O 2 deprivation resulted in a decrease in the isoelectric point of eIF4E, consistent with additional phosphorylation. Modi®cation of eIF4E was mimicked by treatment with caffeine under aerobic conditions and blocked by treatment with ruthenium red under O 2 deprivation, implicating Ca 2+ as a second messenger in eIF4E modi®cation. In contrast, no isoelectric variants of eIFiso4E were detected. The possible role of cytosolic Ca 2+ and pH in regulation of mRNA cap-binding protein activity under O 2 deprivation is discussed.

Phosphoglucomutase (PGM) catalyzes the interconversion of glucose (Glc)-1-and Glc-6-phosphate in the synthesis and consumption of sucrose. We isolated two maize (Zea mays L.) cDNAs that encode PGM with 98.5% identity in their deduced amino acid sequence. Southern-blot analysis with genomic DNA from lines with different Pgm1 and Pgm2 genotypes suggested that the cDNAs encode the two known cytosolic PGM isozymes, PGM1 and PGM2. The cytosolic PGMs of maize are distinct from a plastidic PGM of spinach (Spinacia oleracea). The deduced amino acid sequences of the cytosolic PGMs contain the conserved phosphate-transfer catalytic center and the metal-ion-binding site of known prokaryotic and eukaryotic PGMs. PGM mRNA was detectable by RNA-blot analysis in all tissues and organs examined except silk. A reduction in PGM mRNA accumulation was detected in roots deprived of O 2 for 24 h, along with reduced synthesis of a PGM identified as a 67-kD phosphoprotein on two-dimensional gels. Therefore, PGM is not one of the so-called "anaerobic polypeptides." Nevertheless, the specific activity of PGM was not significantly affected in roots deprived of O 2 for 24 h. We propose that PGM is a stable protein and that existing levels are sufficient to maintain the flux of Glc-1-phosphate into glycolysis under O 2 deprivation.

Maize cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) is encoded by a small multi-gene family consisting of gpc1, gpc2, gpc3 and gpc4. GAPC3/4 protein is synthesized in roots during anoxic conditions and is known to be one of the 'anaerobic polypeptides'. We further analyzed the gpc gene family by isolating full-length cDNA clones of gpc2, gpc3, gpc4 and genomic clones of gpc2 and gpc4. The deduced amino acid sequence of GAPC4 has 99.4% identity with that of GAPC3 as compared to only 81% with either GAPC1 or GAPC2 amino acid sequence. Based on the deduced amino acid sequence identity we designated GAPC1 and GAPC2 as group I (97% identical) and GAPC3 and GAPC4 as group II (99.4% identical). As previously reported for gpc3, transcript levels were also induced for gpc4 by anaerobiosis. Neither heat shock, cold nor salt stress induced the expression of gpc3 or gpc4. In contrast, the transcript accumulation of gpc1 and gpc2 either remained constitutive or decreased in response to anoxia. The upstream regions of gpc2 and gpc4 contain typical eukaryotic promoter features with transcription start points at 76 and 68 bp upstream of their respective translation initiation sites. Transient expression analysis of gpc4 promoter-beta-glucuronidase (GUS) reporter gene constructs in bombarded maize suspension culture cells was used to examine the role of 5'-flanking sequence of gpc4. The gpc4 promoter (-1997 to +39 bp) was sufficient to induce GUS activity approximately three-fold in response to anaerobiosis. 5'-unidirectional deletion analysis revealed that the critical region of gpc4 required for its induced expression lies between -290 and -157. This region has reverse-oriented putative 'anaerobic response elements', G-box like sequences, and a GC motif similar to that previously defined as a regulatory element of maize adh1 and Arabidopsis adh, as well as the sequences found in other environmentally inducible genes. The relevance of these elements in conferring anaerobic induction of gpc4 gene expression is discussed.

BACKGROUND: Tuberculosis is considered to be a major cause of death among the infectious diseases prevalent in India. Differentiation of M. tuberculosis from Nontuberculous Mycobacteria (NTM) is very important in the management of the disease. Recently, Standard Diagnostics (SD Yongin Korea) developed a simple and rapid assay using a mouse monoclonal anti-MPT64 antibody to discriminate between M. tuberculosis and NTM by Immunochromatography. These tests were found to have good sensitivity and specificity. METHOD: 200 sputum samples were processed and inoculated on conventional Lowenstein Jensen culture medium. The Mycobacterial isolates were subjected to biochemical tests (Niacin test and Nitrate reduction test) and Immunochromatographic assay. The isolates were also tested by Polymerase chain reaction. H37 RV strain was used as a positive control. RESULT: The culture isolates showed positive in 47 (23.5%) cases. 44 isolates were positive and 3 were negative by both biochemical tests and immunochromatographic assay. Similar results were obtained by PCR. The sensitivity and specificity of Immunochromatography test when compared to biochemical tests and PCR was 100%. Positive predictive value and Negative predictive values were 100%. CONCLUSION: Immunochromatographic assay showed comparable results with biochemical tests and this assay can be used as an alternative to biochemical tests for the confirmation of Mycobacterium tuberculosis complex, which avoids use of hazardous chemicals (Cyanogen bromide).