			Home About us Contact

Proteolytic Cascade (proteolytic + cascade)

Distribution by Scientific Domains

Life Sciences	80%

Selected Abstracts

Inhibition of urokinase receptor gene expression and cell invasion by anti-uPAR DNAzymes in osteosarcoma cells

FEBS JOURNAL, Issue 14 2005
Charles E. De Bock
The urokinase-type plasminogen activator (uPA) receptor (uPAR) has been implicated in signal transduction and biological processes including cancer metastasis, angiogenesis, cell migration, and wound healing. It is a specific cell surface receptor for its ligand uPA, which catalyzes the formation of plasmin from plasminogen, thereby activating the proteolytic cascade that contributes to the breakdown of extracellular matrix, a key step in cancer metastasis. We have synthesized three different DNA enzymes (Dz372, Dz483 and Dz720) targeting uPAR mRNA at three separate purine (A or G),pyrimidine (U or C) junctions. Two of these DNAzymes, Dz483 and Dz720, cleaved uPAR transcript in vitro with high efficacy and specificity at a molar ratio (uPAR to Dz) as low as 1 : 0.2. When analyzed over 2 h with a 200-fold molar excess of DNAzymes to uPAR transcript, Dz720 and Dz483 were able to decrease uPAR transcript in vitro by ,,93% and ,,84%, respectively. They also showed an ability to cleave uPAR mRNA in the human osteosarcoma cell line Saos-2 after transfection. The DNAzyme Dz720 decreased uPAR mRNA within 4 h of transfection, and inhibited uPAR protein concentrations by 55% in Saos-2 cells. The decrease in uPAR mRNA and protein concentrations caused by Dz720 significantly suppressed Saos-2 cell invasion as assessed by an in vitro Matrigel assay. The use of DNAzyme methodology adds a new potential clinical agent for decreasing uPAR mRNA expression and inhibiting cancer invasion and metastasis. [source]

Caspase cleavage of exon 9 deleted presenilin-1 is an early event in apoptosis induced by calcium ionophore A 23187 in SH-SY5Y neuroblastoma cells

JOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2001
Bogdan O. Popescu
Abstract Presenilins (PSs) are mutated in a majority of familial Alzheimer disease (FAD) cases. Mutated PSs may cause FAD by a number of pro-apoptotic mechanisms, or by regulating ,-secretase activity, a protease involved in ,-amyloid precursor protein processing to the neurotoxic ,-amyloid peptide. Besides their normal endoproteolytic processing, PSs are substrates for caspases, being cleaved to alternative N-terminal and C-terminal fragments. So far little is known about the role of PSs cleavage in the apoptotic machinery. Here, we used SH-SY5Y neuroblastoma cells stably transfected with wild-type or exon 9 deleted presenilin 1 (PS1) in a time-course study after the exposure to the calcium ionophore A23187. During and after exposure to A 23187, intracellular calcium levels were higher in exon 9 deleted PS1 cells as compared with non-transfected and wild-type PS1 transfected cells. Cell death and the enrichment of apoptotic cells after A23187 exposure were increased by overexpression of exon 9 deleted PS1 as compared with the control cell lines. Wild-type PS1 cells were compared with exon 9 deleted PS1 cells and the temporal relationship between PS1 and other caspase substrates cleavages was analyzed. Exon 9 deleted PS1 cells exhibited a higher caspase-3 activation and a greater cleavage of PS1 and poly(ADP-ribose) polymerase (PARP) compared with wild-type PS1 cells. Exon 9 deleted PS1 cleavage occurred earlier than other caspase substrate cleavages (i.e., PARP and gelsolin), simultaneous with minimum detectable caspase-3 activation. Therefore, alternative cleavage of PS1 may play an important role for the regulation of the proteolytic cascade activated during apoptosis. J. Neurosci. Res. 66:122,134, 2001. © 2001 Wiley-Liss, Inc. [source]

Rapid autocatalytic activation of the M4 metalloprotease aureolysin is controlled by a conserved N-terminal fungalysin-thermolysin-propeptide domain

MOLECULAR MICROBIOLOGY, Issue 6 2008
Nicholas N. Nickerson
Summary The Staphylococcus aureus proteolytic cascade consists of a metalloprotease aureolysin (Aur), which activates a serine protease zymogen proSspA, which in turn activates the SspB cysteine protease. As with other M4 metalloproteases, including elastase of Pseudomonas aeruginosa, the propeptide of proAur contains an N-terminal fungalysin-thermolysin-propeptide (FTP) domain. Autocatalytic activation of proAur was initiated by processing at T85,L86 in the FTP domain. This differed from the mechanism described for proElastase, where the FTP domain has an RY motif in place of TL86, and processing occurred at the junction of the propeptide and metalloprotease domains, which remained as an inactive complex during passage across the outer membrane. When TL86 in the FTP domain was replaced with RY, an intact N-terminal propeptide was secreted, but the M4 metalloprotease domain was degraded. Consequently, this segment of the FTP domain promotes intramolecular processing of proAur while bestowing a chaperone function, but discourages processing within the FTP domain of proElastase, where activation must be co-ordinated with passage across a second membrane. We conclude that the FTP domain of proAur is adapted to facilitate a rapid autocatalytic activation mechanism, consistent with the role or proAur as initiator of the staphylococcal proteolytic cascade. [source]

Serine proteases from two cell types target different components of a complex that governs regulated intramembrane proteolysis of pro-,K during Bacillus subtilis development

MOLECULAR MICROBIOLOGY, Issue 3 2005
Ruanbao Zhou
Summary Upon starvation Bacillus subtilis undergoes a developmental process involving creation of two cell types, the mother cell and forespore. A signal in the form of a serine protease, SpoIVB, is secreted from the forespore and leads to regulated intramembrane proteolysis (RIP) of pro-,K, releasing active ,K into the mother cell. RIP of pro-,K is carried out by a membrane-embedded metalloprotease, SpoIVFB, which is inactive when bound by BofA and SpoIVFA. We have investigated the mechanism by which this complex is activated. By expressing components of the signalling pathway in Escherichia coli, we reconstructed complete inhibition of pro-,K RIP by BofA and SpoIVFA, and found that SpoIVB serine protease activity could partially restore RIP, apparently by targeting SpoIVFA. Pulse-chase experiments demonstrated that SpoIVFA synthesized early during B. subtilis sporulation is lost in a SpoIVB-dependent fashion, coincident with the onset of pro-,K RIP, supporting the idea that SpoIVB targets SpoIVFA to trigger RIP of pro-,K. Loss of BofA depended not only on SpoIVB, but also on CtpB, a serine protease secreted from the mother cell. CtpB appeared to cleave BofA near its C-terminus upon coexpression in E. coli, and purified CtpB degraded BofA. We propose that RIP of pro-,K involves a three-step proteolytic cascade in which SpoIVB first cleaves SpoIVFA, CtpB then cleaves BofA and finally SpoIVFB cleaves pro-,K. [source]