MIP Genes are Down-regulated Under Drought Stress in Nicotiana glauca

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Abstract
Water flux across cell membranes has been shown to occur not only through the lipid bilayer, but also through aquaporins, which are members of the major intrinsic protein (MIP) super-family of channel proteins. Aquaporins greatly increase the membrane permeability for water, but may also be regulated, allowing cellular control over the rate of water influx/efflux. Water flux is crucial for stomatal opening and closing, but little is known about the role that aquaporins play in stomatal physiology. Our initial goal was to isolate and characterize the MIP genes expressed in guard cells of the model plant, Nicotiana glauca. Degenerate oligonucleotides corresponding to amino acid sequences conserved in tonoplast intrinsic proteins (TIPs) or plasma membrane intrinsic proteins (PIPs) were used to amplify portions of MIP genes by RT-PCR. These PCR products were used as probes in screening a N. glauca guard cell cDNA library. We isolated three clones (NgMIP1, NgMIP2 and NgMIP3) homologous to TIPs and two clones (NgMIP4 and NgMIP5) homologous to PIPs. All of the MIP genes we characterized displayed highest levels of mRNA accumulation in roots or stems, with lower levels of expression in mesophyll cells and whole leaves, and lowest transcript accumulation in guard cell RNA. Interestingly, the accumulation of transcripts arising from NgMIP2, NgMIP3 and NgMIP4 diminished dramatically in drought-stressed plants. This down-regulation of MIP gene expression may result in reduced membrane water permeability and may encourage cellular water conservation during periods of dehydration stress.