In recent years, members of the protein kinase family have been discovered at an accelerated pace. Most were first described, not through the traditional biochemical approach of protein purification and enzyme assay, but as putative protein kinase amino acid sequences deduced from the nucleotide sequences of molecularly cloned genes or complementary DNAs. Phylogenetic mapping of the conserved protein kinase catalytic domains can serve as a useful first step in the functional characterization of these newly identified family members.

http://science.sciencemag.org/content/sci/241/4861/42.full.pdf

The protein kinase family: conserved features and deduced phylogeny of the catalytic domains.

The capacity of the adult brain and spinal cord to repair lesions by axonal regeneration or compensatory fibre growth is extremely limited. A monoclonal antibody (IN-1) raised against NI-220/250, a myelin protein that is a potent inhibitor of neurite growth, promoted axonal regeneration and compensatory plasticity following lesions of the central nervous system (CNS) in adult rats. Here we report the cloning of nogo A, the rat complementary DNA encoding NI-220/250. The nogo gene encodes at least three major protein products (Nogo-A, -B and -C). Recombinant Nogo-A is recognized by monoclonal antibody IN-1, and it inhibits neurite outgrowth from dorsal root ganglia and spreading of 3T3 fibroblasts in an IN-1-sensitive manner. Antibodies against Nogo-A stain CNS myelin and oligodendrocytes and allow dorsal root ganglion neurites to grow on CNS myelin and into optic nerve explants. These data show that Nogo-A is a potent inhibitor of neurite growth and an IN-1 antigen produced by oligodendrocytes, and may allow the generation of new reagents to enhance CNS regeneration and plasticity.

Nogo-A is a myelin-associated neurite outgrowth inhibitor and an antigen for monoclonal antibody IN-1

Publisher Summary Early studies pointed to the directions for subsequent investigations and there has been continuing identification of other circulating autoantibodies, characterization of their respective cellular antigens, and demonstration of the relationship between autoantibodies and clinical syndromes. The antibodies identified in these studies have been used extensively by investigators in molecular and cell biology as powerful probes for understanding the precursor messenger RNA (mRNA) splicing. Autoantibodies designated SS-A or Ro and SS-B or La were identified in patients with Sjogren's syndrome (SS) and SLE, and have been shown to target intracellular proteins that may be involved with regulation of RNA polymerase III function. Investigators in the field had long perceived that these spontaneously occurring auto-antibodies in human disease would turn out to be useful reagents in cell biology. It was also appreciated that it was necessary to characterize the molecular nature of the auto-antigens so that in addition to the immunochemical properties, their structure and function might be determined. One of the purposes of this chapter is to show that the new molecular biology of cellular antigens and auto-antibodies could now be providing insights into comprehending some features of autoimmunity. In addition, there appears to be a need for the synthesis of the wealth of information that has accumulated and an evaluation of what it might signify.

Antinuclear antibodies: diagnostic markers for autoimmune diseases and probes for cell biology

A Multibasic Cleavage Site in the Spike Protein of SARS-CoV-2 Is Essential for Infection of Human Lung Cells.

The present invention is directed to secreted and transmembrane polypeptides and to nucleic acid molecules encoding those polypeptides. Also provided herein are vectors and host cells comprising those nucleic acid sequences, chimeric polypeptide molecules comprising the polypeptides of the present invention fused to heterologous polypeptide sequences, antibodies which bind to the polypeptides of the present invention and to methods for producing the polypeptides of the present invention.

Secreted and Transmembrane Polypeptides and Nucleic Acids Encoding the Same

We have examined the role of Furin in postimplantation-stage mouse embryos by analyzing both the expression pattern of fur mRNA and the developmental consequences of a loss-of-function mutation at the fur locus. At early stages (day 7.5), fur mRNA is abundant in extraembryonic endoderm and mesoderm, anterior visceral endoderm, and in precardiac mesoderm. 1 day later fur is expressed throughout the heart tube and in the lateral plate mesoderm, notochordal plate and definitive gut endoderm. Embryos lacking Furin die between days 10.5 and 11.5, presumably due to hemodynamic insufficiency associated with severe ventral closure defects and the failure of the heart tube to fuse and undergo looping morphogenesis. Morphogenesis of the yolk sac vasculature is also abnormal, although blood islands and endothelial precursors form. Analysis of cardiac and endodermal marker genes shows that while both myocardial precursors and definitive endoderm cells are specified, their numbers and migratory properties are compromised. Notably, mutant embryos fail to undergo axial rotation, even though Nodal and eHand, two molecular markers of left-right asymmetry, are appropriately expressed. Overall, the present data identify Furin as an important activator of signals responsible for ventral closure and embryonic turning.

Failure of ventral closure and axial rotation in embryos lacking the proprotein convertase furin

The human fur gene encodes a protein, designated furin, the C-terminal half of which contains a transmembrane and a cysteine-rich receptor-like domain. The N-terminal half of furin exhibits striking primary amino acid sequence similarity to the catalytic domains of members of the subtilisin family of serine proteases. We here report characteristics of the furin protein and propose a three-dimensional model for its presumptive catalytic domain with characteristics, that predict furin to exhibit an endoproteolytic cleavage selectivity at paired basic residues. This prediction is substantiated by transfection and cotransfection experiments, using COS-1 cells. Full length fur cDNA evokes the specific synthesis of two polypeptides of about 100 kDa and 90 kDa as appeared from Western blot analysis of transfected COS-1 cells using a polyclonal anti-furin antiserum. Functional analysis of furin was performed by cotransfection of fur cDNA with cDNA encoding the 'wild type' precursor of von Willebrand factor (pro-vWF) and revealed an increased proteolytic processing of provWF. In contrast, cotransfection of fur cDNA with a recombinant derivative (provWFgly763), having the arginine residue adjacent to the proteolytic cleavage site (arg-ser-lys-arg) replaced by glycine, revealed that provWFgly763 is not processed by the fur gene product. We conclude that in higher eukaryotes, furin is the prototype of a subtilisin-like class of proprotein processing enzymes with substrate specificity for paired basic residues.

Furin is a subtilisin-like proprotein processing enzyme in higher eukaryotes

Recently we described that genetic sequences in the immediately upstream region of the c-fes/fps proto-oncogene, designated fur, constituted a transcription unit for a 45-kb mRNA Here we present characteristics of the genetic organization of fur and some features of its putative translation product which we call furin The nucleotide sequence of a 31-kbp fur-specific cDNA isolated from a human cDNA library revealed an open reading frame of 1,498 bp from which the 499 carboxy-terminal amino acids of the primary fur translational product could be deduced Computer analysis indicated that furin contained a possible transmembrane domain which resembled that of class II MHC antigens Furthermore, a cysteine-rich region was present Significant homology, especially with respect to the topography of cysteine residues, was found between the cysteine-rich regions of the human insulin receptor, the human epidermal growth factor receptor and furin From the data presented here we deduce that fur may encode a membrane-associated protein with a recognition function

Evolutionary conserved close linkage of the c-fes/fps proto-oncogene and genetic sequences encoding a receptor-like protein.

The recently discovered fur gene encodes a membrane-associated protein with a recognition function. To further characterize the gene, we studied its expression by Northern blot analysis using poly(A)-selected RNA from a variety of organs of African green monkey and rat. The fur gene appeared to be differentially expressed, relatively high levels of fur mRNA being present in specimens of liver and kidney, low levels in brain, spleen, and thymus, and very low levels in heart muscle, lung, and testis. mRNA levels in specimens of human lung tissue without neoplastic lesions were also very low. Similar analyses of primary human lung carcinomas of different histopathological types revealed a highly selective and strong elevation of fur expression in nonsmall cell lung carcinomas, but not in small cell lung carcinomas. These results indicate that fur expression can be used to discriminate between these two types of human lung cancer.

/pdf/fur-gene-expression-as-a-discriminating-marker-for-small-re80eutr5d.pdf

A. J. M. Roebroek

Papers

Failure of ventral closure and axial rotation in embryos lacking the proprotein convertase furin

Furin is a subtilisin-like proprotein processing enzyme in higher eukaryotes

Evolutionary conserved close linkage of the c-fes/fps proto-oncogene and genetic sequences encoding a receptor-like protein.

fur gene expression as a discriminating marker for small cell and nonsmall cell lung carcinomas.

Modulation of furin-mediated proprotein processing activity by site-directed mutagenesis