Phototropins are blue-light (BL) receptor serine (Ser)/threonine kinases, and contain two light, oxygen, and voltage (LOV) domains, and are members of the PAS domain superfamily. They mediate phototropism, chloroplast movement, leaf expansion, and stomatal opening of higher plants in response to BL. In stomatal guard cells, genetic analysis has revealed that phototropins mediate activation of the plasma membrane H ϩ -ATPase by phosphorylation and drive stomatal opening. However, biochemical evidence for the involvement of phototropins in the BL response of stomata is lacking. Using guard cell protoplasts, we showed that broad bean (Vicia faba) phototropins (Vfphots) were phosphorylated by BL, and that this phosphorylation of Vfphots reached to the maximum level earlier than that of the H ϩ -ATPase. Phosphorylation of both Vfphots and H ϩ -ATPase showed similar sensitivity to BL and were similarly suppressed by protein kinase and flavoprotein inhibitors. We found that a 14-3-3 protein was bound to Vfphots upon phosphorylation, and this binding occurred earlier than the H ϩ -ATPase phosphorylation. Vfphots (Vfphot1a and Vfphot1b) were expressed in Escherichia coli, and phosphorylation sites were determined to be Ser-358 for Vfphot1a and Ser-344 for Vfphot1b, which are localized between LOV1 and LOV2. We conclude that Vfphots act as BL receptors in guard cells and that phosphorylation of a Ser residue between LOV1 and LOV2 and subsequent 14-3-3 protein binding are likely to be key steps of BL response in stomata. The binding of a 14-3-3 protein to Vfphot was found in etiolated seedlings and leaves in response to BL, suggesting that this event was common to phototropin-mediated responses.Stomatal pores surrounded by a pair of guard cells in the epidermis regulate gas exchange between leaves and the atmosphere and allow CO 2 entry for both photosynthesis and the transpirational stream in higher plants (Zeiger, 1983; Assmann, 1993). Stomata open through the activation of a H ϩ pump in guard cells in response to blue light (BL; Assmann et al., 1985; Shimazaki et al., 1986). The BL-activated pump creates an inside-negative, electrical potential across the plasma membrane and drives K ϩ uptake through voltage-gated K ϩ channels (Hedrich and Schroeder, 1989; Assmann and Shimazaki, 1999; Schroeder et al., 2001). The H ϩ pump has been demonstrated to be the plasma membrane H ϩ -ATPase and is activated via phosphorylation of its C terminus with concomitant binding of the 14-3-3 protein (Kinoshita and Shimazaki, 1999; Emi et al., 2001; Palmgren, 2001). Although physiological responses downstream of BL perception have been elucidated extensively in guard cells, the mechanism of BL perception itself, an initial event in the BL response of stomata, had yet to be elucidated. Recently, genetic analysis of Arabidopsis mutants strongly suggests that phototropins (Atphot1 and Atphot2) function as BL receptors in a redundant manner and mediate BL-dependent stomatal opening (Kinoshita et al., 2001), although the biochemical eviden...

Phototropins (phot1 and phot2) are suggested to be multifunctional blue-light (BL) receptors mediating phototropism, chloroplast movement, stomatal opening, and leaf expansion. The Arabidpsis phot1 phot2 double mutant lacks all of these responses. To confirm the requirement of phototropins in BL responses, the Arabidopsis phot1 phot2 double mutant was transformed with PHOT1 cDNA and the phenotypic restoration was analysed in the transformants. It was found that all BL responses were restored, although differentially, by the transformation of the Arabidopsis phot1 phot2 double mutant with PHOT1 cDNA. The results showed that phot1 was an essential component for all these BL responses in planta, and that the cellular level of phot1 might determine the individual BL responses.

We analysed spatial patterns of expression of a lacZ reporter gene in the gut of Drosophila larvae that had been transformed with a P-element-lacZ vector to identify regional differences in gene expression. lacZ-positive epithelial cells formed distinct domains with discrete transverse and longitudinal boundaries along the gut tube. Boundaries were often found at sites at which morphological boundaries were not obvious. The gut epithelium was subdivided into 36 compartments by the boundaries. We refer to these novel compartments as "tissue compartments". The lacZ-positive domain of each strain appeared as a single tissue compartment or as a combination of several tissue compartments. The tissue compartment is considered to be a unit of regional differentiation. The spatial organization of the tissue compartments may represent the "floor plan", determined by genes that control the regional differentiation of this nonsegmental organ.

The proctodeum of the Drosophila embryo originates from the posterior end of the blastoderm and forms the hindgut. By enhancer-trap mutagenesis, using a P-element-lacZ vector, we identified a mutation that caused degeneration of the proctodeum during shortening of the germ band and named it aproctous (apro). Expression of the lacZ reporter gene, which was assumed to represent expression of the apro gene, began at the cellular blastoderm stage in a ring that encompassed about 10-15% of the egg's length (EL) and included the future proctodeum, anal pads, and posterior-most part of the visceral mesoderm. In later stages, strong expression of lacZ was detected in the developing hindgut and anal pads. Expression continued in the anal pads and epithelium of the hindgut of larvae; the epithelium of the hindgut of the adult fly also expressed lacZ. The spatial patterns of the expression of lacZ in various mutants suggested that the embryonic expression of apro was regulated predominantly by two gap genes, tailless (tll) and huckebein (hkb): tll is necessary for the activation of apro, while hkb suppressed the expression of apro in the region posterior to 10% EL. Cloning and sequencing of the apro cDNA revealed that apro was identical to the T-related gene (Trg) that is a Drosophila homolog of the vertebrate Brachyury gene. apro appears to play a key role in the development of tissues derived from the proctodeum.