Regulation of phytochrome B signaling by phytochrome A and FHY1 in Arabidopsis thaliana

Phytochrome A (phyA) and phytochrome B (phyB) share the control of many processes but little is known about mutual signaling regulation. Here, we report on the interactions between phyA and phyB in the control of the activity of an Lhcb1*2 gene fused to a reporter, hypocotyl growth and cotyledon unfolding in etiolated Arabidopsis thaliana. The very-low fluence responses (VLFR) induced by pulsed far-red light and the high-irradiance responses (HIR) observed under continuous far-red light were absent in the phyA and phyA phyB mutants, normal in the phyB mutant, and reduced in the fhy1 mutant that is defective in phyA signaling. VLFR were also impaired in Columbia compared to Landsberg erecta. The low-fluence responses (LFR) induced by red-light pulses and reversed by subsequent far-red light pulses were small in the wild type, absent in phyB and phyA phyB mutants but strong in the phyA and fhy1 mutants. This indicates a negative effect of phyA and FHY1 on phyB-mediated responses. However, a pre-treatment with continuous far-red light enhanced the LFR induced by a subsequent red-light pulse. This enhancement was absent in phyA, phyB, or phyA phyB and partial in fhy1. The levels of phyB were not affected by the phyA or fhy1 mutations or by far-red light pre-treatments. We conclude that phyA acting in the VLFR mode (i.e. under light pulses) is antagonistic to phyB signaling whereas phyA acting in the HIR mode (i.e. under continuous far-red light) operates synergistically with phyB signaling, and that both types of interaction require FHY1.     

The extremely conserved C-terminal region of Reelin is not necessary for secretion but is required for efficient activation of downstream signaling

Reelin is a very large secreted glycoprotein essential for correct development of the mammalian brain. It is also implicated in higher functions and diseases of human brain. However, whether or not secretion of Reelin is regulated and how Reelin transmits signals remain largely unknown. Reelin protein is composed of an N-terminal F-spondin-like domain, Reelin repeats, and a short and highly basic C-terminal region (CTR). The primary sequence of CTR is almost completely conserved among vertebrates except fishes, indicating its importance. A prevailing idea regarding the function of CTR is that it is required for the secretion of Reelin, although this remains unproven. Here we aimed to clarify the function of Reelin CTR. Neither deleting most of CTR nor replacing CTR with unrelated amino acids affected secretion efficiency, indicating that CTR is not absolutely required for the secretion of Reelin. We also found that Reelin mutants without CTR were less potent in activating the downstream signaling in cortical neurons. Although these mutants were able to bind to the Reelin receptor ectodomain as efficiently as wild-type Reelin, quite interestingly, their ability to bind to the isolated cell membrane bearing Reelin receptors or receptor-expressing cells (including cortical neurons) was much weaker than that of wild-type Reelin. Therefore, it is concluded that the CTR of Reelin is not essential for its secretion but is required for efficient activation of downstream signaling events, presumably via binding to an unidentified "co-receptor" molecule(s) on the cell membrane.     

Calcium is sufficient but not necessary for activation of sheep platelet cytosolic phospholipase A2

In this study we demonstrate that: (1) although the major phospholipase A2 present in sheep platelets is activated by calcium ions, it can effectively catalyze hydrolysis of the sn-2 ester linkage in phospholipids in the absence of calcium; (2) expression of calcium-independent phospholipase A2 activity can be induced by NaCl utilizing purified (but not crude) cytosolic enzyme; and (3) calcium-independent phospholipase A2 activity is regulated by a reconstitutable cytosolic protein. Collectively, these results underscore the fundamental catalytic differences between extracellular and intracellular calcium-dependent phospholipases A2 and demonstrate that calcium is sufficient, but not necessary, for the activation of this class of intracellular phospholipases A2.     

Deglycosylation is necessary but not sufficient for activation of proconcanavalin A.

Concanavalin A (ConA), one of the most studied plant lectins, is formed in jack bean (Canavalia ensiformis) seeds. ConA is synthesized as an inactive glycoprotein precursor proConA. Different processing events such as endoproteolytic cleavages, ligation of peptides and deglycosylation of the precursor are required to generate the different polypeptides constitutive of mature ConA. Among these events, deglycosylation of the prolectin appears as a key step in the lectin activation. The detection of deglycosylated proConA in immature jack bean seeds indicates that endoproteolytic cleavages are not prerequisite for its deglycosylation. Both the structure of the lectin precursor N-glycans Man8-9GlcNAc2 and the capacity of Endo H to cleave these oligosaccharide from native proConA in vitro favoured Endo H-type glycosidases as candidates for proConA deglycosylation in planta. Evidence for pH-dependent changes in the prolectin folding were obtained from analysis of the N-glycan accessibility and activation of the deglycosylated lectin precursor in acidic conditions. These data are consistent with the observation that both deglycosylation and acidification of the pH are the minimum requirements to convert the inactive precursor into an active lectin.     

Repression of c-myc is necessary but not sufficient for terminal differentiation of B lymphocytes in vitro

The importance of c-myc as a target of the Blimp-1 repressor has been studied in BCL-1 cells, in which Blimp-1 is sufficient to trigger terminal B-cell differentiation. Our data show that Blimp-1-dependent repression of c-myc is required for BCL-1 differentiation, since constitutive expression of c-Myc blocked differentiation. Furthermore, ectopic expression of cyclin E mimicked the effects of c-Myc on both proliferation and differentiation, indicating that the ability of c-Myc to drive proliferation is responsible for blocking BCL-1 differentiation. However, inhibition of c-Myc by a dominant negative form was not sufficient to drive BCL-1 differentiation. Thus, during Blimp-1-dependent plasma cell differentiation, repression of c-myc is necessary but not sufficient, demonstrating the existence of additional Blimp-1 target genes.     

TAB2 is essential for prevention of apoptosis in fetal liver but not for interleukin-1 signaling

The proinflammatory cytokine interleukin-1 (IL-1) transmits a signal via several critical cytoplasmic proteins such as MyD88, IRAKs and TRAF6. Recently, serine/threonine kinase TAK1 and TAK1 binding protein 1 and 2 (TAB1/2) have been identified as molecules involved in IL-1-induced TRAF6-mediated activation of AP-1 and NF-kappa B via mitogen-activated protein (MAP) kinases and I kappa B kinases, respectively. However, their physiological functions remain to be clarified. To elucidate their roles in vivo, we generated TAB2-deficient mice. The TAB2 deficiency was embryonic lethal due to liver degeneration and apoptosis. This phenotype was similar to that of NF-kappa B p65-, IKK beta-, and NEMO/IKK gamma-deficient mice. However, the IL-1-induced activation of NF-kappa B and MAP kinases was not impaired in TAB2-deficient embryonic fibroblasts. These findings demonstrate that TAB2 is essential for embryonic development through prevention of liver apoptosis but not for the IL-1 receptor-mediated signaling pathway.     

Preferential role for NF-kappa B/Rel signaling in the type 1 but not type 2 T cell-dependent immune response in vivo.

T cell function is a critical determinant of immune responses as well as susceptibility to allergic diseases. Activated T cells can differentiate into effectors whose cytokine profile is limited to type 1 (IFN-gamma-dominant) or type 2 (IL-4-, IL-5-dominant) patterns. To investigate mechanisms that connect extracellular stimuli with the regulation of effector T cell function, we have measured immune responses of transgenic mice whose NF-kappa B/Rel signaling pathway is inhibited in T cells. Surprisingly, these mice developed type 2 T cell-dependent responses (IgE and eosinophil recruitment) in a model of allergic pulmonary inflammation. In contrast, type 1 T cell responses were severely impaired, as evidenced by markedly diminished delayed-type hypersensitivity responses, IFN-gamma production, and Ag-specific IgG2a levels. Taken together, these data indicate that inhibition of NF-kappa B can lead to preferential impairment of type 1 as compared with type 2 T cell-dependent responses.     

Destruction of Xenopus cyclins A and B2, but not B1, requires binding to p34cdc2.

The specific and rapid destruction of cyclins A and B during mitosis is their most remarkable property. A short peptide motif of approximately 10 amino acids near the N-terminus, known as the destruction box, is absolutely required for programmed proteolysis. In this paper we show that although the destruction box is necessary for the degradation of cyclin A, it is not sufficient. Mutant versions of cyclin A that cannot form complexes with p34cdc2 are stable, which we interpret to mean that this cyclin must bind to p34cdc2 in order to undergo programmed proteolysis. Thus, N-terminal fragments of cyclin A containing little more than the destruction box and its surroundings are indestructible. p34cdc2 binding also appears to be required for the destruction of cyclin B2. In contrast, cyclin B1 does not require p34cdc2 binding for specific proteolysis. The systems for the proteolysis of cyclins A, B1 and B2 thus appear to show important differences in the way they recognize their substrates.