RESUMO
SignificanceClassic serine proteases are synthesized as inactive precursors that are proteolytically processed, resulting in irreversible activation. We report an alternative and reversible mechanism of activation that is executed by an inactive protease. This mechanism involves a protein complex between the serine protease HTRA1 and the cysteine protease calpain 2. Surprisingly, activation is restricted as it improves the proteolysis of soluble tau protein but not the dissociation and degradation of its amyloid fibrils, a task that free HTRA1 is efficiently performing. These data exemplify a challenge for protein quality control proteases in the clearing of pathogenic fibrils and suggest a potential for unexpected side effects of chemical modulators targeting PDZ or other domains located at a distance to the active site.
Assuntos
Calpaína , Serina Endopeptidases , Amiloide/metabolismo , Calpaína/metabolismo , Serina Peptidase 1 de Requerimento de Alta Temperatura A/química , Proteólise , Serina Endopeptidases/metabolismo , Serina Proteases/metabolismoRESUMO
Communication between plant cells and their biotic environment largely depends on the function of plasma membrane localized receptor-like kinases (RLKs). Major players in this communication within root meristems are secreted peptides, including CLAVATA3/EMBRYO SURROUNDING REGION40 (CLE40). In the distal root meristem, CLE40 acts through the RLK ARABIDOPSIS CRINKLY4 (ACR4) and the leucine-rich repeat (LRR) RLK CLAVATA1 (CLV1) to promote cell differentiation. In the proximal meristem, CLE40 signaling requires the LRR receptor-like protein CLAVATA2 (CLV2) and the membrane localized pseudokinase CORYNE (CRN) and serves to inhibit cell differentiation. The molecular components that act immediately downstream of the CLE40-activated receptors are not yet known. Here, we show that active CLE40 signaling triggers the release of intracellular Ca2+ leading to increased cytosolic Ca2+ concentration ([Ca2+]cyt) in a small subset of proximal root meristem cells. This rise in [Ca2+]cyt depends on the CYCLIC NUCLEOTIDE GATED CHANNELS (CNGCs) 6 and 9 and on CLV1. The precise function of changes in [Ca2+]cyt is not yet known but might form a central part of a fine-tuned response to CLE40 peptide that serves to integrate root meristem growth with stem cell fate decisions and initiation of lateral root primordia.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cálcio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Canais de Cátion Regulados por Nucleotídeos Cíclicos/metabolismo , Meristema/metabolismo , Raízes de Plantas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Diferenciação Celular/genética , Canais de Cátion Regulados por Nucleotídeos Cíclicos/genética , Variação Genética , Genótipo , Meristema/genética , Raízes de Plantas/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
A significant part of the communication between plant cells is mediated by signaling peptides and their corresponding plasma membrane-localized receptor-like kinases. This communication mechanism serves as a key regulatory unit for coordination of plant growth and development. In the past years more peptide-receptor signaling pathways have been shown to regulate developmental processes, such as shoot and root meristem maintenance, seed formation, and floral abscission. More detailed understanding of the processes behind this regulation might also be helpful to increase the yield of crop plants.
Assuntos
Peptídeos/metabolismo , Desenvolvimento Vegetal , Transdução de Sinais , Agricultura , Modelos Biológicos , Plantas/metabolismo , Plantas/microbiologia , Processamento de Proteína Pós-TraducionalRESUMO
Mammalian high-temperature requirement serine protease A1 (HTRA1) is a secreted member of the trypsin family of serine proteases which can degrade a variety of bone matrix proteins and as such has been implicated in musculoskeletal development. In this study, we have investigated the role of HTRA1 in mesenchymal stem cell (MSC) osteogenesis and suggest a potential mechanism through which it controls matrix mineralization by differentiating bone-forming cells. Osteogenic induction resulted in a significant elevation in the expression and secretion of HTRA1 in MSCs isolated from human bone marrow-derived MSCs (hBMSCs), mouse adipose-derived stromal cells (mASCs), and mouse embryonic stem cells. Recombinant HTRA1 enhanced the osteogenesis of hBMSCs as evidenced by significant changes in several osteogenic markers including integrin-binding sialoprotein (IBSP), bone morphogenetic protein 5 (BMP5), and sclerostin, and promoted matrix mineralization in differentiating bone-forming osteoblasts. These stimulatory effects were not observed with proteolytically inactive HTRA1 and were abolished by small interfering RNA against HTRA1. Moreover, loss of HTRA1 function resulted in enhanced adipogenesis of hBMSCs. HTRA1 Immunofluorescence studies showed colocalization of HTRA1 with IBSP protein in osteogenic mASC spheroid cultures and was confirmed as being a newly identified HTRA1 substrate in cell cultures and in proteolytic enzyme assays. A role for HTRA1 in bone regeneration in vivo was also alluded to in bone fracture repair studies where HTRA1 was found localized predominantly to areas of new bone formation in association with IBSP. These data therefore implicate HTRA1 as having a central role in osteogenesis through modification of proteins within the extracellular matrix.
