Lactacystin-induced apoptosis of cultured mouse cortical neurons is associated with accumulation of PTEN in the detergent-resistant membrane fraction

The tumor suppressor function of PTEN is attributed to its phospholipid phosphatase activity that dephosphorylates the plasma membrane phosphatidylinositol-(3,4,5)-triphosphate [PtdIns(3,4,5)P₃]. Implicit in this notion is that PTEN needs to be targeted to the plasma membrane to dephosphorylate PtdIns(3,4,5)P₃. However, the recruitment of PTEN to the plasma membrane is not fully understood. Here, we demonstrate PTEN accumulation in the detergent-insoluble fraction of neuronal cells in response to treatment by the proteasome inhibitor lactacystin. First, lactacystin induces apoptosis and the activation of caspase-3 in cultured cortical neurons. Second, PTEN undergoes proteolysis to form a truncated 50-kDa form that lacks parts of its C-terminal tail. Third, the truncated PTEN is stably associated with the detergent-insoluble fraction in which the plasma membrane marker protein flotillin-1 resides. Taken together, our results suggest that truncation and accumulation of PTEN to the detergent-insoluble membrane fraction are two events associated with the apoptotic signals of the proteasome inhibitor in cortical neurons.Received 24 March 2004; received after revision 26 May 2004; accepted 5 June 2004     

Synthesis and properties of crosslinked recombinant pro-resilin

Resilin is a member of a family of elastic proteins that includes elastin, as well as gluten, gliadin, abductin and spider silks. Resilin is found in specialized regions of the cuticle of most insects, providing low stiffness, high strain and efficient energy storage; it is best known for its roles in insect flight and the remarkable jumping ability of fleas and spittle bugs. Previously, the Drosophila melanogaster CG15920 gene was tentatively identified as one encoding a resilin-like protein (pro-resilin). Here we report the cloning and expression of the first exon of the Drosophila CG15920 gene as a soluble protein in Escherichia coli. We show that this recombinant protein can be cast into a rubber-like biomaterial by rapid photochemical crosslinking. This observation validates the role of the putative elastic repeat motif in resilin function. The resilience (recovery after deformation) of crosslinked recombinant resilin was found to exceed that of unfilled synthetic polybutadiene, a high resilience rubber. We believe that our work will greatly facilitate structural investigations into the functional properties of resilin and shed light on more general aspects of the structure of elastomeric proteins. In addition, the ability to rapidly cast samples of this biomaterial may enable its use in situ for both industrial and biomedical applications.     

Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus

Spike (S) proteins of coronaviruses, including the coronavirus that causes severe acute respiratory syndrome (SARS), associate with cellular receptors to mediate infection of their target cells. Here we identify a metallopeptidase, angiotensin-converting enzyme 2 (ACE2), isolated from SARS coronavirus (SARS-CoV)-permissive Vero E6 cells, that efficiently binds the S1 domain of the SARS-CoV S protein. We found that a soluble form of ACE2, but not of the related enzyme ACE1, blocked association of the S1 domain with Vero E6 cells. 293T cells transfected with ACE2, but not those transfected with human immunodeficiency virus-1 receptors, formed multinucleated syncytia with cells expressing S protein. Furthermore, SARS-CoV replicated efficiently on ACE2-transfected but not mock-transfected 293T cells. Finally, anti-ACE2 but not anti-ACE1 antibody blocked viral replication on Vero E6 cells. Together our data indicate that ACE2 is a functional receptor for SARS-CoV.     

Biologically active <Emphasis Type="Italic">cis</Emphasis>-cinnamic acid occurs naturally in <Emphasis Type="Italic">Brassica parachinensis</Emphasis>

The biologically active cis-cinnamic acid (cis-CA) has been perceived as a synthetic plant growth regulator for decades,However,in the present study,we found that cis-CA actually exists as a naturally occurring compound in a Brassica plant,This natural growth-regulating substance presents in both the sunlight-irradiated leaf tissue and the non-irradiated root tissue ,The concentrations of cis-CA in both tissues are comparable to the bilogi-cally effective lvels of those major plant hormones,the presence of cis-CA in root tissue suggests that it may be produced through both light-dependent and -independent path-ways or it can be transproted from a plant organ to another.     

Enzymes for chemical synthesis

New catalytic synthetic methods in organic chemistry that satisfy increasingly stringent environmental constraints are in great demand by the pharmaceutical and chemical industries. In addition, novel catalytic procedures are necessary to produce the emerging classes of organic compounds that are becoming the targets of molecular and biomedical research. Enzyme-catalysed chemical transformations are now widely recognized as practical alternatives to traditional (non-biological) organic synthesis, and as convenient solutions to certain intractable synthetic problems.     

New developments in enzymatic peptide synthesis.

This review article describes new enzymatic methods developed for the efficient and irreversible synthesis of peptides based on native and modified proteases, and for the synthesis of polypeptides containing D- and/or unnatural amino acids. Potential opportunities for future developments in the field based on new enzymes, tailor-made catalytic antibodies, and on the technique of in vitro mutagenesis are also described.     

Single neurones can initiate synchronized population discharge in the hippocampus

The synchronized firing of neuronal populations is frequently observed in the mammalian central nervous system. The generation of motor activities such as locomotion and respiration requires the simultaneous activation of many neurones and synchronous firing also underlies the cortical alpha rhythm and the hippocampal theta rhythm. However the influence that single neurones may have on such neuronal population discharges is not clear. We have examined this question using small isolated segments of the CA3 region of the guinea pig hippocampus. We report here that in the presence of picrotoxin, a gamma-aminobutyric acid (GABA) antagonist, these segments spontaneously generate synchronized rhythmic bursts comparable with the interictal epileptiform discharges observed in the hippocampus and neocortex in the presence of penicillin. The activation of some individual neurones by intracellular current injection can partially entrain and reset the rhythm. The probability that a synchronized burst will follow stimulation of a single cell increases with time after a spontaneous synchronized discharge, suggesting that each population discharge is followed by a period of relative population refractoriness. A delay of 40-200 ms elapses between the activation of a single neurone and the synchronized discharge. We suggest that during this time activity elicited in one neurone spreads to other neurones through multisynaptic excitatory pathways and leads eventually to the participation of the whole population in a synchronous burst.