TOPCONS (http://topcons.net/) is a widely used web server for consensus prediction of membrane protein topology. We hereby present a major update to the server, with some substantial improvements, ...including the following: (i) TOPCONS can now efficiently separate signal peptides from transmembrane regions. (ii) The server can now differentiate more successfully between globular and membrane proteins. (iii) The server now is even slightly faster, although a much larger database is used to generate the multiple sequence alignments. For most proteins, the final prediction is produced in a matter of seconds. (iv) The user-friendly interface is retained, with the additional feature of submitting batch files and accessing the server programmatically using standard interfaces, making it thus ideal for proteome-wide analyses. Indicatively, the user can now scan the entire human proteome in a few days. (v) For proteins with homology to a known 3D structure, the homology-inferred topology is also displayed. (vi) Finally, the combination of methods currently implemented achieves an overall increase in performance by 4% as compared to the currently available best-scoring methods and TOPCONS is the only method that can identify signal peptides and still maintain a state-of-the-art performance in topology predictions.
Cathepsin B (CTSB) is a powerful lysosomal protease. This review evaluated
gene knockout (KO) outcomes for amelioration of brain dysfunctions in neurologic diseases and aging animal models. Deletion ...of the
gene resulted in significant improvements in behavioral deficits, neuropathology, and/or biomarkers in traumatic brain injury, ischemia, inflammatory pain, opiate tolerance, epilepsy, aging, transgenic Alzheimer's disease (AD), and periodontitis AD models as shown in 12 studies. One study found beneficial effects for double
and cathepsin S KO mice in a multiple sclerosis model. Transgenic AD models using amyloid precursor protein (APP) mimicking common sporadic AD in three studies showed that
KO improved memory, neuropathology, and biomarkers; two studies used APP representing rare familial AD and found no
KO effect, and two studies used highly engineered APP constructs and reported slight increases in a biomarker. In clinical studies, all reports found that CTSB enzyme was upregulated in diverse neurologic disorders, including AD in which elevated CTSB was positively correlated with cognitive dysfunction. In a wide range of neurologic animal models, CTSB was also upregulated and not downregulated. Further, human genetic mutation data provided precedence for CTSB upregulation causing disease. Thus, the consilience of data is that
gene KO results in improved brain dysfunction and reduced pathology through blockade of CTSB enzyme upregulation that causes human neurologic disease phenotypes. The overall findings provide strong support for CTSB as a rational drug target and for CTSB inhibitors as therapeutic candidates for a wide range of neurologic disorders. SIGNIFICANCE STATEMENT: This review provides a comprehensive compilation of the extensive data on the effects of deleting the cathepsin B (
) gene in neurological and aging mouse models of brain disorders. Mice lacking the
gene display improved neurobehavioral deficits, reduced neuropathology, and amelioration of neuronal cell death and inflammatory biomarkers. The significance of the compelling CTSB evidence is that the data consilience validates CTSB as a drug target for discovery of CTSB inhibitors as potential therapeutics for treating numerous neurological diseases.
During normal aging, innate immunity progresses to a chronic state. However, how oxidative stress and chronic neuroinflammation arise during aging remains unclear. In this study, we found that ...genetic ablation of cathepsin B (CatB) in mice significantly reduced the generation of reactive oxygen species (ROS) and neuroinflammation and improved cognitive impairment during aging. In cultured microglia, pharmacological inhibition of CatB significantly reduced the generation of mitochondria‐derived ROS and proinflammatory mediators induced by L‐leucyl‐L‐leucine methyl ester (LLOMe), a lysosome‐destabilizing agent. In the CatB‐overexpressing microglia after treatment with LLOMe, which mimicked the aged microglia, CatB leaked in the cytosol is responsible for the degradation of the mitochondrial transcription factor A (TFAM), resulting in the increased generation of mitochondria‐derived ROS and proinflammatory mediators through impaired mtDNA biosynthesis. Furthermore, intralateral ventricle injection of LLOMe‐treated CatB‐overexpressing microglia induced cognitive impairment in middle‐aged mice. These results suggest that the increase and leakage of CatB in microglia during aging are responsible for the increased generation of mitochondria‐derived ROS and proinflammatory mediators, culminating in memory impairment.
Abstract
Cathepsin D (CTSD) is a lysosomal protease and a marker of poor prognosis in breast cancer. However, the cells responsible for this association and the function of CTSD in cancer are still ...incompletely understood. By using a conditional CTSD knockout mouse crossed to the transgenic
MMTV-PyMT
breast cancer model we demonstrate that CTSD deficiency in the mammary epithelium, but not in myeloid cells, blocked tumor development in a cell-autonomous manner. We show that lack of CTSD impaired mechanistic Target of Rapamycin Complex 1 (mTORC1) signaling and induced reversible cellular quiescence. In line, CTSD-deficient tumors started to grow with a two-month delay and quiescent
Ctsd
-/-
tumor cells re-started proliferation upon long-term culture. This was accompanied by rewiring of oncogenic gene expression and signaling pathways, while mTORC1 signaling remained permanently disabled in CTSD-deficient cells. Together, these studies reveal a tumor cell-autonomous effect of CTSD deficiency, and establish a pivotal role of this protease in the cellular response to oncogenic stimuli.
The general view on cysteine cathepsins, which were long believed to be primarily involved in intracellular protein turnover, has dramatically changed in last 10 to 15 years. The discovery of new ...cathepsins, such as cathepsins K, V, X, F and O, and their tissue distribution suggested that at least some of them are involved in very specific cellular processes. Moreover, gene ablation experiments revealed that cathepsins play a vital role in numerous physiological processes, such as antigen processing and presentation, bone remodelling, prohormone processing and wound healing. Their involvement in several pathologies, including osteoporosis, rheumatoid arthritis, osteoarthritis, bronchial asthma and cancer have also been confirmed and today several of them have been validated as relevant targets for therapies. Compounds targeting cathepsins S and K are already in clinical evaluation, whereas others are in experimental phases. The cathepsin K inhibitor AAE-581 (balicatib) as the most advanced of them passed Phase II clinical trials in 2005. In this review, we discuss the current view on cathepsins as an emerging group of targets for several diseases and the development of cathepsin K and S inhibitors for treatment of osteoporosis and various immune disorders.
Cathepsin K is responsible for the degradation of type I collagen in osteoclast-mediated bone resorption. Collagen fragments are known to be biologically active in a number of cell types. Here, we ...investigate their potential to regulate osteoclast activity. Mature murine osteoclasts were seeded on type I collagen for actin ring assays or dentine discs for resorption assays. Cells were treated with cathepsins K-, L-, or MMP-1-predigested type I collagen or soluble bone fragments for 24 h. The presence of actin rings was determined fluorescently by staining for actin. We found that the percentage of osteoclasts displaying actin rings and the area of resorbed dentine decreased significantly on addition of cathepsin K-digested type I collagen or bone fragments, but not with cathepsin L or MMP-1 digests. Counterintuitively, actin ring formation was found to decrease in the presence of the cysteine proteinase inhibitor LHVS and in cathepsin K-deficient osteoclasts. However, cathepsin L deficiency or the general MMP inhibitor GM6001 had no effect on the presence of actin rings. Predigestion of the collagen matrix with cathepsin K, but not by cathepsin L or MMP-1 resulted in an increased actin ring presence in cathepsin K-deficient osteoclasts. These studies suggest that cathepsin K interaction with type I collagen is required for 1) the release of cryptic Arg-Gly-Asp motifs during the initial attachment of osteoclasts and 2) termination of resorption via the creation of autocrine signals originating from type I collagen degradation.
Interleukin (IL)-1β and IL-18 play critical roles in the induction of chronic pain hypersensitivity. Their inactive forms are activated by caspase-1. However, little is known about the mechanism ...underlying the activation of pro-caspase-1. There is increasing evidence that cathepsin B (CatB), a typical lysosomal cysteine protease, is involved in the pro-caspase-1 activation and the subsequent maturation of IL-1β and IL-18. In this context, CatB is considered to be an important molecular target to control chronic pain. However, no information is currently available about the role of CatB in chronic pain hypersensitivity. We herein show that CatB deficiency or the intrathecal administration of CA-074Me, a specific CatB inhibitor, significantly inhibited the induction of complete Freund's adjuvant-induced tactile allodynia in mice without affecting peripheral inflammation. In contrast, CatB deficiency did not affect the nerve injury-induced tactile allodynia. Furthermore, CatB deficiency or CA-074Me treatment significantly inhibited the maturation and secretion of IL-1β and IL-18 by cultured microglia following treatment with the neuroactive glycoprotein chromogranin A (CGA), but not with ATP. Moreover, the IL-1β expression in spinal microglia and the induction of tactile allodynia following the intrathecal administration of CGA depended on CatB, whereas those induced by the intrathecal administration of ATP or lysophosphatidic acid were CatB independent. These results strongly suggest that CatB is an essential enzyme for the induction of chronic inflammatory pain through its activation of pro-caspase-1, which subsequently induces the maturation and secretion of IL-1β and IL-18 by spinal microglia. Therefore, CatB-specific inhibitors may represent a useful new strategy for treating inflammation-associated pain.
Zusammenfassung
Visitationen in Kirchengemeinden prägen seit Jahrhunderten die Führung der reformatorischen
Kirche. Ursprünglich aus einem eher direktiven Ansatz der Prüfung
und Kontrolle ...hervorgegangen, fließen in den letzten Jahren auch stärker beraterische
und kooperative Ideen in dieses Instrument ein. Anhand einer empirischen Studie in
vier deutschen EKD-Gliedkirchen wurde die Visitation bezüglich ihrer Steuerungswirkung
untersucht. Das besondere Augenmerk galt dabei der Frage, wie Visitationen
die Gliederung und Verschiedenheit des kirchlichen Lebens aufgreifen und in die
Gesamtkirche integrieren können. Mit einer theoretischen Vorklärung werden einige
organisations- und kirchensoziologische Aspekte zusammengetragen.
Abstract
For centuries, visitations have shaped leadership and governance in the Protestant
church. Historically, the visitation is based on a rather directive approach focusing on
leadership and control; more recently, it has been propagated as a cooperative instrument
that allows for consulting and organizational development during the visitation
process. Using empirical data from four territorial churches – “Landeskirchen”
– in Germany, this article explores the usage and functions of the visitation as governance
instrument. We pay particular attention to the question how the visitation
addresses the varying contexts and loosely coupled structures in the different church
communities and how it helps integrating the single communities into the overall
Protestant church. Selected concepts from organization sociology and religious organizations
are used as a theoretical framework and interpretation scheme.
The translocon recognizes sufficiently hydrophobic regions of a protein and inserts them into the membrane. Computational methods try to determine what hydrophobic regions are recognized by the ...translocon. Although these predictions are quite accurate, many methods still fail to distinguish marginally hydrophobic transmembrane (TM) helices and equally hydrophobic regions in soluble protein domains. In vivo, this problem is most likely avoided by targeting of the TM-proteins, so that non-TM proteins never see the translocon. Proteins are targeted to the translocon by an N-terminal signal peptide. The targeting is also aided by the fact that the N-terminal helix is more hydrophobic than other TM-helices. In addition, we also recently found that the C-terminal helix is more hydrophobic than central helices. This information has not been used in earlier topology predictors.
Here, we use the fact that the N- and C-terminal helices are more hydrophobic to develop a new version of the first-principle-based topology predictor, SCAMPI. The new predictor has two main advantages; first, it can be used to efficiently separate membrane and non-membrane proteins directly without the use of an extra prefilter, and second it shows improved performance for predicting the topology of membrane proteins that contain large non-membrane domains.
The predictor, a web server and all datasets are available at http://scampi.bioinfo.se/
arne@bioinfo.se
Supplementary data are available at Bioinformatics online.
Multiple types of degradative enzymes, including cathepsins of the cysteine protease family, have been implicated in the regulation of angiogenesis and invasion during cancer progression. Several ...cysteine cathepsins are up-regulated in a mouse model of pancreatic islet cell carcinogenesis (RIP1-Tag2), and tumor progression is impaired following their collective pharmacologic inhibition. Using null mutations of four of the implicated cysteine cathepsins, we have now dissected their individual roles in cancer development. Mutants of cathepsins B or S impaired tumor formation and angiogenesis, while cathepsin B or L knockouts retarded cell proliferation and tumor growth. Absence of any one of these three genes impaired tumor invasion. In contrast, removal of cathepsin C had no effect on either tumor formation or progression. We have identified E-cadherin as a target substrate of cathepsins B, L, and S, but not cathepsin C, potentially explaining their differential effects on tumor invasion. Furthermore, we detected analogous increases in cathepsin expression in human pancreatic endocrine neoplasms, and a significant association between increased levels of cathepsins B and L and tumor malignancy. Thus individual cysteine cathepsin genes make distinctive contributions to tumorigenesis.
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