Plant pathogen effectors can recruit the host post-translational machinery to mediate their post-translational modification (PTM) and regulate their activity to facilitate parasitism, but few studies have focused on this phenomenon in the field of plant-parasitic nematodes. In this study, we show that the plant-parasitic nematode Meloidogyne graminicola has evolved a novel effector, MgGPP, that is exclusively expressed within the nematode subventral esophageal gland cells and up-regulated in the early parasitic stage of M. graminicola. The effector MgGPP plays a role in nematode parasitism. Transgenic rice lines expressing MgGPP become significantly more susceptible to M. graminicola infection than wild-type control plants, and conversely, in planta, the silencing of MgGPP through RNAi technology substantially increases the resistance of rice to M. graminicola. Significantly, we show that MgGPP is secreted into host plants and targeted to the ER, where the N-glycosylation and C-terminal proteolysis of MgGPP occur. C-terminal proteolysis promotes MgGPP to leave the ER, after which it is transported to the nucleus. In addition, N-glycosylation of MgGPP is required for suppressing the host response. The research data provide an intriguing example of in planta glycosylation in concert with proteolysis of a pathogen effector, which depict a novel mechanism by which parasitic nematodes could subjugate plant immunity and promote parasitism and may present a promising target for developing new strategies against nematode infections.

Tissue factor (TF) is the protein cofactor for factor VIIa (FVIIa), the first serine protease of the clotting cascade. Previous studies using alanine mutagenesis have identified TF residues Lys165 and Lys166 as important for factor X (FX) activation, hypothesizing either that these residues interact with phospholipid head groups or that they directly or indirectly promote macromolecular substrate binding. In the recently reported x-ray crystal structure of the isolated extracellular domain of TF, both Lys165 and Lys166 are solvent-exposed and predicted to be near the phospholipid surface in intact TF. We hypothesized that these residues may in fact be ideally positioned to interact with the 4-carboxyglutamate-rich domain (Gla domain) of FX. We therefore predicted that mutations at Lys165 and Lys166 should have no effect on the activation of Gla domainless FX. To test this hypothesis, we mutated both residues Lys165 and Lys166 of TF to Ala, Glu, or Gln and examined the ability of these double mutants to support FVIIa-mediated activation of FX, Gla domainless FX, and factor IX (FIX). Each TF mutant was equivalent to wild-type TF in both FVIIa binding and promotion of FVIIa amidolytic activity. However, all three mutants were markedly deficient in supporting FIX and FX activation, with FX activation rates decreased more than FIX activation rates. In both reactions, the TF mutants exhibited different extents of activity: Gln165-Gln166 > Ala165-Ala166 > Glu165-Glu166. In sharp contrast, all three TF mutants were equivalent to wild-type TF in supporting activation of Gla domainless FX by FVIIa. Interestingly, the deficiency of the mutants in FX activation was less pronounced when Gla domainless FVIIa was used in place of native FVIIa. Together, these findings suggest that TF residues Lys165 and Lys166 contribute to a binding site for the Gla domain of FX (and perhaps other substrates) and that this interaction may be facilitated by the presence of the Gla domain of FVIIa.

Previous animal studies have demonstrated that alpha-melanocyte-stimulating hormone (alpha-MSH) is a sebotropic hormone in rats and that targeted disruption of melanocortin 5 receptor (MC5-R) can down-regulate sebum output in mice. To study the role of proopiomelanocortin (POMC) peptides in the regulation of human sebaceous lipid production and sebocyte differentiation, we established a primary human sebocyte culture system. Sebocytes were derived from normal human facial skin. Differentiation of sebocytes, induced by POMC-derived peptides such as MSH, adrenocorticotropic hormone (ACTH), or bovine pituitary extract (BPE), resulted in the appearance of prominent cytoplasmic lipid droplets. Partial induction of sebocyte differentiation also was observed in serum-depleted cultures, but there was very limited spontaneous differentiation in serum-containing medium. Analysis by high-performance thin-layer chromatography (HPTLC) of (14)C-acetate-labeled lipids showed a dose-dependent increase in synthesis of sebaceous-specific lipid (i.e., squalene) induced by NDP alpha-MSH. Molecular studies using RT-PCR showed a low level of human MC5-R expression under serum-free condition but a substantial increase after treatment with NDP alpha-MSH or BPE. In contrast, MC1-R expression remained the same, independent of treatment. Our data indicate that expression of MC5-R correlates with sebocyte differentiation and suggest a regulatory role for MC5-R in human sebaceous lipid production.

Overall structure and function of TF-FVlla (1,18,19); it confers the ability to bind reversibly to anionic phospholipids. Next to the Gla-domain, and sometimes classified as part of it, is a short shetch of largely hydrophobic residues called the hydrophobic or aromatic stack. Next are two modules related to epidermal growth factor (EGF domains). EGFr, the more N-terminal of the two, contains a