Cotton annexin proteins participate in the establishment of fiber cell elongation scaffold

Cotton plant is one of the most important economic crops in the world which supplies natural fiber for textile industry. The crucial traits of cotton fiber quality are fiber length and strength, which are mostly determined by the fiber elongation stage. Annexins are assumed to be involved in regulating fiber elongation, but direct evidences remain elusive. Recently, we have investigated the activities of fiber-specific expressed annexins AnGb5/6 and their interacted proteins in cotton. AnGb5 and 6 can interact reciprocally to generate a protein macro-raft in cell membrane. This macro-raft is probably a stabilized scaffold for Actin1 organization. The actin assembling direction and density are correlated with AnGb6 gene expression and fiber expanding rate among three fiber length genotypes. These results suggest that annexins may act as the adaptor that linked fiber cell membrane to actin assembling. Due to the strong Ca²⁺ and lipid binding ability of annexins, these results also indicate that annexins complex may function as an intermediate to receive Ca²⁺ or lipid signals during fiber elongation.     

组蛋白激活拟南芥CDPK催化CaMBP10的体外磷酸化反应

植物转脂蛋白 (plant lipid transfer proteins, LTPs) 是高等植物中广泛存在的多基因编码的小分子碱性蛋白. 本研究室已经证明白菜和豌豆LTPs可分别被内源胞浆可溶性和膜结合钙依赖性蛋白激酶 (calcium-dependent protein kinase, CDPK) 磷酸化. 为深入研究CDPK对白菜钙调素结合蛋白10 (calmodulin-binding protein-10, CaMBP10) 的磷酸化性质及特征, 本文从拟南芥可溶性蛋白粗提物中检测到1个分子量约为54 kD的CDPK对CaMBP10有磷酸化作用. 研究表明, 组蛋白可增强 CDPK对CaMBP10的磷酸化活性, 促进磷酸化进程. 而且组蛋白和Ca2+对CDPK具有协同调节效应, 二者共同作用时比Ca2+单独作用时, 激酶的活力增强约12倍. 此外, 不同组蛋白对CDPK的激活能力不同, 组蛋白1对该激酶活性的激活能力要比组蛋白3高约8倍.     

OsDi19‐4 acts downstream of OsCDPK14 to positively regulate ABA response in rice

The drought‐induced 19 protein family consists of several atypical Cys2/His2‐type zinc finger proteins in plants and plays an important role in abiotic stress. In this study, we found that overexpressing OsDi19‐4 in rice altered the expression of a series of abscisic acid (ABA)‐responsive genes, resulting in strong ABA‐hypersensitive phenotypes including ABA‐induced seed germination inhibition, early seedling growth inhibition and stomatal closure. On the contrary, OsDi19‐4 knockdown lines were less sensitive to ABA. Additionally, OsCDPK14 was identified to interact with OsDi19‐4 and be responsible for the phosphorylation of OsDi19‐4, and the phosphorylation of OsDi19‐4 was further enhanced after the treatment of ABA. Apart from these, OsDi19‐4 was shown to directly bind to the promoters of OsASPG1 and OsNAC18 genes, two ABA‐responsive genes, and regulate their expression. Transient expression assays confirmed the direct regulation role of OsDi19‐4, and the regulation was further enhanced by the increased phosphorylation of OsDi19‐4 after the treatment of ABA. Taken together, these data demonstrate that OsDi19‐4 acts downstream of OsCDPK14 to positively regulate ABA response by modulating the expression of ABA‐responsive genes in rice.     

Phosphorylation of sucrose synthase at serine 170: occurrence and possible role as a signal for proteolysis

Sequence analysis identified serine 170 (S170) of the maize (Zea mays L.) SUS1 sucrose synthase (SUS) protein as a possible, second phosphorylation site. Maize leaves contained two calcium-dependent protein kinase activities and a calcium-independent kinase activity with characteristics of an sucrose non-fermenting 1 (SNF1)-related protein kinase. Phosphorylation of the novel S170 and the known serine 15 (S15) site by these protein kinases was determined in peptide substrates and detected in SUS1 protein substrates utilizing sequence- and phosphorylation-specific antibodies. We demonstrate phosphorylation of S170 in vitro and in vivo. The calcium-dependent protein kinases phosphorylated both S170 and S15, whereas SNF1-related protein kinase activity was restricted to S15. Calcium-dependent protein-kinase-mediated S170 and S15 phosphorylation kinetics were determined in wild-type and mutant SUS1 substrates. These analyses revealed that kinase specificity for S170 was threefold lower than that for S15, and that phosphorylation of S170 was stimulated by prior phosphorylation at the S15 site. The SUS-binding peptides encoded by early nodulin 40 (ENOD40) specifically antagonized S170 phosphorylation in vitro. A model wherein S170 phosphorylation functions as part of a mechanism targeting SUS for proteasome-mediated degradation is supported by the observations that SUS proteolytic fragments: (i) were detected and possessed relatively high phosphorylated-S170 (pS170) stoichiometry; (ii) were spatially coincident with proteasome activity within developing leaves; and (iii) co-sedimented with proteasome activity. In addition, full-length pS170-SUS protein was less stable than S170-SUS in cultured leaf segments and was stabilized by proteasome inhibition. Post-translational control of SUS protein level through pS170-promoted proteolysis may explain the specific and significant decrease in SUS abundance that accompanies the sink-to-source transition in developing maize leaves.     

CPK12: A Ca2+-dependent protein kinase balancer in abscisic acid signaling

Ca²⁺ is believed to be a critical second messenger in ABA signal transduction. Ca²⁺-dependent protein kinases (CDPKs) are the best characterized Ca²⁺ sensors in plants. Recently, we identified an Arabidopsis CDPK member CPK12 as a negative regulator of ABA signaling in seed germination and post-germination growth, which reveals that different members of the CDPK family may constitute a regulation loop by functioning positively and negatively in ABA signal transduction. We observed that both RNA interference and overexpression of CPK12 gene resulted in ABA-hypersensitive phenotypes in seed germination and post-germination growth, suggesting a high complexity of the CPK12-mediated ABA signaling pathway. CPK12 stimulates a negative ABA-signaling regulator (ABI2) and phosphorylates two positive ABA-signaling regulators (ABF1 and ABF4), which may partly explain the ABA hypersensitivity induced by both downregulation and upregulation of CPK12 expression. Our data indicate that CPK12 appears to function as a balancer in ABA signal transduction in Arabidopsis.     

VfCPK1, a gene encoding calcium-dependent protein kinase from Vicia faba, is induced by drought and abscisic acid

Calcium, one of the most ubiquitous second messengers, has been shown to be involved in a wide variety of responses in plants. Calcium-dependent protein kinases (CDPKs) (EC 2.7.1.37) are the predominant Ca(2+)-regulated serine/threonine protein kinase in plants and play an important role in plant calcium signal transduction. CDPKs are encoded by a large multigene family in many plants, which has been showed so far; however, the precise role of each specific CDPK is still largely unknown. A novel CDPK gene designated as VfCPK1 was cloned from epidermal peels of broad bean (Vicia faba L.) leaves using the rapid amplification of cDNA ends (RACE)-PCR technique and its expression was studied in detail. The VfCPK1 cDNA is 1783 bp long and contains an open reading frame of 1482 bp encoding 493 amino acids. VfCPK1 contains all conserved regions found in CDPKs and shows a high level of sequence similarity to many other plant CDPKs. VfCPK1 was highly expressed in leaves, especially in leaf epidermal peels of broad bean in mRNA and protein levels. Expressions of VfCPK1 at both the mRNA and protein levels were increased in leaves treated with abscisic acid or subjected to drought stress. Potential roles of VfCPK1 in epidermal peels are discussed. The nucleotide sequence data reported here were deposited in the GenBank database under accession number AY753552.