;Aquaporins in the plasma and vacuolar membranes play a key role in the intercellular and intracellular water transport in plants. First, we quantitated the absolute amounts for mRNAs of eight aquaporin isoforms in hypocotyls of radish seedlings. Then, we investigated the effects of salt and water stresses (150 mM NaCl, 300 mM mannitol and 20% polyethylene glycol) and phytohormones (gibberellic acid, abscisic acid and brassinolide) on the mRNA and protein levels of aquaporins in the plasma membrane (RsPIP1-1, 1-2, 1-3, 2-1, 2-2 and 2-3) and vacuolar membrane (RsTIP1-1 and 2-1). The mRNA and protein levels of RsTIP1-1, RsTIP2-1, RsPIP1-1, RsPIP1-2 and RsPIP1-3 were comparatively constant. In contrast, mannitol treatment altered the mRNA levels of RsPIP2-1, RsPIP2-2 and RsPIP2-3 in roots. Immunoblot analysis showed that the RsPIP2-1 protein level was increased by NaCl treatment and decreased by treatment with mannitol and polyethylene glycol. Gibberellic acid and abscisic acid suppressed the levels of mRNAs of RsPIP2-1, RsPIP2-2 and RsPIP2-3 and the protein level of RsPIP2-1 in roots. On the other hand, the protein levels of RsPIP1-group members and RsTIPs were scarcely changed by these phytohormones. In the case of hypocotyls and cotyledons, the mRNA and protein levels of eight isoforms were not markedly affected by any treatment. These results indicate that aquaporins in the root, especially the RsPIP2 group, may be a stress responsive type of aquaporin at least in the protein level.

Human sperm nucleoproteins consist of protamines and histones. Changes in composition of these proteins are thought to correlate with spermatogenesis and may be involved in some instances of male infertility. We sought to separate sperm nucleoproteins including variants of protamine using an improved two-dimensional electrophoretic method, with the aim of comprehensively analysing all sperm nucleoprotein constituents. After extracting nuclear basic proteins from the sperm of normal volunteers, we analysed these proteins on a gel sheet by a radical free, highly reducing method based on Kaltschmidt and Whittmann's two-dimensional electrophoresis. Basic proteins from sperm nuclei were separated clearly into 12 spots. By amino acid sequence analysis, these spots corresponded to protamine 1 (P1)- (five spots), protamine 2 (P2)-related proteins (six spots) and testis-specific histone H2B (one spot). The N-terminal amino acid sequences of the six P2-related proteins were compatible with those of HPI1, HPI2, HPS1, HPS2, HP2 and HP3, and quantitative comparison could be performed. In conclusion, human sperm nucleoproteins including all P2-related variants could be analysed quantitatively with high resolution on a single electrophoretic gel.

Previous studies revealed that the recessive allele of the W2 locus generated purple-blue color and high vacuolar pH of flower petals in soybean. The location of W2 gene was reportedly close to simple sequence repeat marker Satt318 in molecular linkage group B2. We used information from the soybean genome to clone a candidate gene for W2. An MYB transcription factor gene belonging to G20 group was found in the vicinity of Satt318. Full-length cDNAs were cloned from purple-flowered cultivar Harosoy (W2 allele) and purple-blue flowered cultivars, Nezumisaya and w2-20 (w2 allele), by reverse transcription-PCR and designated as GmMYB-G20-1. Its open reading frame was 1083 bp long that encoded 361 amino acids in Harosoy. GmMYB-G20-1 had 53.7% similarity in amino acid sequence with the PH4 gene of petunia controlling blueness and vacuolar pH of flower petals. GmMYB-G20-1 of Nezumisaya and w2-20 had 3 base substitutions compared with that of Harosoy. The first substitution generated a stop codon in the MYB domain, resulting in truncated polypeptides. Cleaved amplified polymorphic sequence (CAPS) marker was developed to detect the base substitution. The polymorphic CAPS marker co-segregated with alleles at the W2 locus in the F(2) population. These results suggest that GmMYB-G20-1 might correspond to the W2 gene.