Project description:Proteins from "pressure-loving" piezophiles appear to adapt by greater compressibility via larger total cavity volume. However, larger cavities in proteins have been associated with lower unfolding pressures. Here, dihydrofolate reductase (DHFR) from a moderate piezophile Moritella profunda (Mp) isolated at ~2.9 km in depth and from a hyperpiezophile Moritella yayanosii (My) isolated at ~11 km in depth were compared using molecular dynamics simulations. Although previous simulations indicate that MpDHFR is more compressible than a mesophile DHFR, here the average properties and a quasiharmonic analysis indicate that MpDHFR and MyDHFR have similar compressibilities. A cavity analysis also indicates that the three unique mutations in MyDHFR are near cavities, although the cavities are generally similar in size in both. However, while a cleft overlaps an internal cavity, thus forming a pathway from the surface to the interior in MpDHFR, the unique residue Tyr103 found in MyDHFR forms a hydrogen bond with Leu78, and the sidechain separates the cleft from the cavity. Thus, while Moritella DHFR may generally be well suited to high-pressure environments because of their greater compressibility, adaptation for greater depths may be to prevent water entry into the interior cavities.
Project description:Moritella viscosa is a Gram-negative pathogen that causes large, chronic ulcers, known as winter-ulcer disease, in the skin of several fish species including Atlantic salmon. We used a bath challenge approach to profile the transcriptome responses of M. viscosa-infected Atlantic salmon skin at the lesion (Mv-At) and away from the lesion (Mv-Aw) sites. M. viscosa infection was confirmed through RNA-based qPCR assays. RNA-Seq identified 5212 and 2911 transcripts differentially expressed in the Mv-At compared to no-infection control and Mv-Aw groups, respectively. Also, there were 563 differentially expressed transcripts when comparing the Mv-Aw to control samples. Our results suggest that M. viscosa caused massive and strong, but largely infection site-focused, transcriptome dysregulations in Atlantic salmon skin, and its effects beyond the skin lesion site were comparably subtle. The M. viscosa-induced transcripts of Atlantic salmon were mainly involved in innate and adaptive immune response-related pathways, whereas the suppressed transcripts by this pathogen were largely connected to developmental and cellular processes. As validated by qPCR, M. viscosa dysregulated transcripts encoding receptors, signal transducers, transcription factors and immune effectors playing roles in TLR- and IFN-dependent pathways as well as immunoregulation, antigen presentation and T-cell development. This study broadened the current understanding of molecular pathways underlying M. viscosa-triggered responses of Atlantic salmon, and identified biomarkers that may assist to diagnose and combat this pathogen.
Project description:Shotgun proteomic analysis of 3D-grown or on CAM-assay grown lung cancer cells lacking or expressing the major triglyceride lipase adipose triglyceride lipase (ATGL)
Project description:Treatment of rice tissues with purified preparations of a Xanthomonas oryzae pv. oryzae (Xoo) secreted plant cell wall degrading enzyme, Lipase/Esterase (LipA), elicits cell wall damage induced innate immune responses. LipA activity is required for induction of defense responses. In order to characterize the early events during elaboration of cell wall degrading enzyme, Lipase/Esterase (LipA) induced innate immune response in rice, we have performed global gene expression profiling of rice leaves treated with purified LipA at early time points, 30 minutes and 120min, after treatment. Whole genome transcriptional profiling was performed using Affymetrix Rice GeneChips Leaves of two weeks old green house grown susceptible rice cultivar (Taichung Native-1; TN-1) were infiltrated with either 30-40μl of purified Xoo Lipase/Esterase (LipA)(500μg/ml) or with buffer (10mM potassium phosphate buffer pH 6.0) alone (as described in Jha et al. 2007; MPMI vol 20, pp 31-40). The plants were shifted to a growth chamber (28oC; 80% relative humidity; 12/12h light/dark cycle) 48 hours before the treatment. 20-30 leaf pieces covering the infiltrated zone from each of the treatments were harvested 30 min. and 120 min. after infiltration. Total RNA isolated from the pooled samples was subjected for expression analysis using Affymetrix GeneChip System. The experiment was repeated with three different biological replicates using RNA isolated from three batches of rice leaves treated with the freshly purified Xoo Lipase/Esterase (LipA)and the buffer Gene Expression profiling of rice leaves undergoing an innate immune respone induced by LipA (Lipase/Esterase A) enzyme