Interleukin-1 beta (IL-1 beta)-converting enzyme cleaves the IL-1 beta precursor to mature IL-1 beta, an important mediator of inflammation. The identification of the enzyme as a unique cysteine protease and the design of potent peptide aldehyde inhibitors are described. Purification and cloning of the complementary DNA indicates that IL-1 beta-converting enzyme is composed of two nonidentical subunits that are derived from a single proenzyme, possibly by autoproteolysis. Selective inhibition of the enzyme in human blood monocytes blocks production of mature IL-1 beta, indicating that it is a potential therapeutic target.

Cellular toxicity resulting from nucleation-dependent polymerization of amyloid beta-peptide (Abeta) is considered to be a major and possibly the primary component of Alzheimer's disease (AD). Inhibition of Abeta polymerization has thus been identified as a target for the development of therapeutic agents for the treatment of AD. The intrinsic affinity of Abeta for itself suggested that Abeta-specific interactions could be adapted to the development of compounds that would bind to Abeta and prevent it from polymerizing. Abeta-derived peptides of fifteen residues were found to be inhibitory of Abeta polymerization. The activity of these peptides was subsequently enhanced through modification of their amino termini with specific organic reagents. Additional series of compounds prepared to probe structural requirements for activity allowed reduction of the size of the inhibitors and optimization of the Abeta-derived peptide portion to afford a lead compound, cholyl-Leu-Val-Phe-Phe-Ala-OH (PPI-368), with potent polymerization inhibitory activity but limited biochemical stability. The corresponding all-D-amino acyl analogue peptide acid (PPI-433) and amide (PPI-457) retained inhibitory activity and were both stable in monkey cerebrospinal fluid for 24 h.

Cysteine proteases related to mammalian interleukin-1 beta-converting enzyme (ICE) and the nematode cell death abnormal ced-3 gene product have been implicated in the effector mechanism of apoptotic cell death. Two novel members of this new family of ICE/CED-3-related proteases, designated ICErel-II and ICErel-III, were cloned from human monocytic cells. Both were highly homologous to human ICE (52% identical) and CED-3 (25% identical) and both contained the absolutely conserved pentapeptide sequence Gln-Ala-Cys-Arg-Asp containing the catalytic cysteine residue. Other structural motifs that were comparable with ICE suggest that ICErel-II and ICErel-III are also synthesized as larger proenzymes which are proteolytically processed to form heterodimeric active enzymes. Pro-interleukin-1 beta processing activity could not be detected in cells transfected with ICErel-II or ICErel-III, but pro-domain-less truncated forms of ICErel-II and ICErel-III were capable of effectively inducing fibroblast apoptosis. ICErel-II and ICErel-III may, therefore, participate in proteolytic events culminating in the apoptotic death of human cells.

Lentiviruses, as exemplified by visna virus of sheep, are nononcogenic retroviruses that cause slowly progressive diseases after prolonged periods of incubation. Earlier studies on visna have shown that the long incubation period of the disease is associated with constant production of minimal quantities of virus in tissues, whereas virus could be obtained by culturing monocytes and macrophages from explants of lymphatic tissues and inflamed organs. In this study the role of macrophages in lentivirus infection was explored using two sheep that were intrabronchially inoculated with virus.

We have isolated cDNA clones encoding the four different forms of mouse myelin basic protein (MBP) and have analyzed the structure of the MBP gene. The three larger forms of MBP differ from the smallest by the inclusion of either or both of two short amino acid sequences at positions 57 and 124 of the smallest protein. The mouse genome contains a single MBP gene comprised of seven exons. The two amino acid sequences present only in the larger MBPs are encoded by separate exons. Furthermore, all exons in the coding region begin or end in complete codons so that alternative splicing does not alter the reading frame. We conclude that the four forms of this myelin protein are encoded in separate mRNAs, each derived by a simple alternative splicing of the primary MBP gene transcript. Comparison of the amino acid sequence encoded by each exon with a recent model of the secondary structure of MBP suggests that each of the seven exons encodes one or two of the predicted structural motifs of the protein.

Neurons in rat central nervous system (CNS) that express 5-HT1c receptor mRNA have been localized by in situ hybridization histochemistry. The 5-HT1c receptor is expressed in a wide variety of cortical and subcortical neurons including hippocampal pyramidal neurons, neurons within most of the central monoaminergic cell groups, neurons in thalamic sensory relay nuclei, and neurons involved in the central processing and regulation of nociceptive transmission.Therefore, the 5-HTic receptor is a prominent but poorly characterized central subclass of serotonin (5-HT) receptor. The distribution of the 5-HTlc receptor within the CNS is considerably more widespread than that of the structurally and functionally related 5-HT2 receptor.The monoamine serotonin (5-hydroxytryptamine; 5-HT) functions as a neurotransmitter in the mammalian central and peripheral nervous systems (1). Within the brain, serotonergic neurons originate primarily in the raphe nuclei of the brainstem (2). The axons of these neurons project to most areas of the central nervous system (CNS) (3, 4), where they regulate a wide variety of sensory, motor, and cortical functions (5). The diverse actions of 5-HT are mediated by interactions with several distinct subclasses of 5-HT receptors. These multiple 5-HT receptors have been defined on the basis of their ligand-binding profile and include . Moreover, individual 5-HT receptor subtypes interact with distinct guanine nucleotide binding proteins and trigger different second-messenger pathways. Activation of the 5-HTla receptor results in an inhibition of adenylate cyclase, whereas activation of the 5-HT1c and 5-HT2 receptors activates phosphotidylinositol-specific phospholipase C, causing the release of the second messengers 1P3 and diacyl glycerol (9, 10).The diverse physiologic actions of 5-HT may not only derive from the activation of different intracellular signaling pathways but also from the expression of the individual receptor subtypes in distinct regions of the brain. Selective pharmacologic ligands have been used to determine the regional distribution of 5-HT receptors in the mammalian CNS. 5-HTla receptors are widely expressed in cortical and subcortical areas, whereas 5-HT2 receptors are detected at highest levels in the cerebral cortex (11, 12). The 5-HT1c subtype was first detected in choroid plexus epithelial cells (13)(14)(15) and is the only 5-HT receptor subtype expressed by these cells. However, the distribution of 5-HT1c receptors in other regions of the nervous system has been difficult because there are no selective ligands for this receptor subclass; most ligands that bind to 5-HT1c receptor also interact with 5-HT2 receptors.Recently, cDNA clones encoding the 5-HTla, 5-HT1c, and 5-HT2 receptors have been isolated and sequenced (16-18). The deduced amino acid sequences reveal that the 5-HT receptors share sequence and structural similarities with the family of guanine nucleotide-binding protein-coupled receptors thought to traverse the membrane seven times (19). The 5-HT1c an...

Shiverer (shi) is an autosomal recessive mutation in the mouse characterized by an almost total lack of central nervous system myelin. While small amounts of other myelin components are present in the brain of the shi mouse, the four forms of myelin basic protein (MBP) are not detectable. Previous investigations by us and others indicate that the MBP gene has undergone a major rearrangement in the shi mutant. Herein, we report in detail the nature and extent of the rearrangement: a 20-kilobase region within the MBP gene is missing in the mutant. We map the 5' breakpoint ofthe deletion to the second intron and the 3' breakpoint to a site 2 kilobases beyond the last MBP exon. The junction of the upstream and downstream portions of the gene contains only one nucleotide not accounted for by the wild-type MBP gene sequence. The 3' side of the deletion occurs in the 3rd of 11 tandem repeats of a 31-base-pair sequence. This region is rich in alternating purine and pyrimidine stretches, sequences that have been associated with both Z-DNA structures and gene rearrangements. Mammalian oligodendrocytes in the central nervous system (CNS) and Schwann cells in the peripheral nervous system (PNS) produce a highly specialized multilamellar myelin membrane that surrounds the neuronal axons and greatly facilitates conduction of electrical impulses (reviewed in ref. 1). The myelin of the CNS has a relatively simple composition in which a family of closely related myelin basic proteins (MBPs) accounts for 30-40% of the total protein in the sheath (reviewed in ref. 2). Peripheral nervous system myelin contains a smaller and dispensable amount of the same MBPs (3, 4). Recent work by us (5) and others (6) has shown that a single gene encodes the family of MBPs and that the various forms of the MBPs are generated by a mechanism of alternative splicing of the mRNA.A number of mutations have been identified in mice that affect oligodendrocyte function and/or Schwann cell function and, consequently, lead to dysmyelination in the CNS and/or the PNS (reviewed in ref. 7). The shiverer (shi) mutation is autosomal recessive and is characterized by the onset of tremors at about the 12th day of life, seizures at later times, and a progressive deterioration ending in an early death (8, 9). The mutation principally affects oligodendrocyte functions and the CNS of the shi mice is almost entirely devoid of myelin membrane (10). The normal amount of myelin is present in the PNS of these animals, but differences in its structure have been noted (4,11,12). The primary defect in shi mice appears to be the absence of MBP (13). By radioimmune assay, MBP levels in shi mice are <0.1% that of wild type (14).The shi allele has been mapped to chromosome 18 by Sidman et al. (15) using classic genetic techniques. The gene encoding the MBP family has also been mapped to chromosome 18 by in situ hybridization and Southern blot analysis of DNA isolated from mouse-hamster somatic cell hybrids (16). Recent work by Roach et al. (17,18) and by us (5) demonst...

The T4 molecule may serve as a T-cell receptor recognizing molecules on the surface of specific target cells and also serves as the receptor for the human immunodeficiency virus. To define the mechanisms of interaction of T4 with the surface of antigen-presenting cells as well as with human immunodeficiency virus, we have further analyzed the sequence, structure, and expression of the human and mouse T4 genes. T4 consists of an extracellular segment comprised of a leader sequence followed by four tandem variable-joining (VJ)-like domains, a transmembrane domain, and a cytoplasmic segment. The structural domains of the T4 protein deduced from amino acid sequence are precisely reflected in the intron-exon organization of the gene. Analysis of the expression of the T4 gene indicates that T4 RNA is expressed not only in T lymphocytes, but in B cells, macrophages, and granulocytes. T4 is also expressed in a developmentally regulated manner in specific regions of the brain. It is, therefore, possible that T4 plays a more general role in mediating cell recognition events that are not restricted to the cellular immune response.Analysis of the surface glycoproteins of peripheral T lymphocytes demonstrates that mature T cells segregate into one of two classes: those that express the surface glycoprotein T4 (CD4) and those that express the glycoprotein T8 (CD8) (1). The T4 molecule is primarily expressed on helper T lymphocytes, whereas T8 is expressed on cytotoxic and suppressor T cells (2, 3). T8+ T lymphocytes interact with a broad set of target cells that express class I major histocompatibility complex (MHC) gene products whereas T4+ T cells interact with a more restricted subset of targets, largely macrophages and B cells, that express class II MHC molecules (2, 3). This has led to the suggestion that the specificity of interaction of subpopulations of T lymphocytes with various target cells results in part from the association of T4 and T8 with the products of different MHC genes. T4 may not only serve as a receptor recognizing molecules on the surface of target cells, but also serves as the receptor for the human immunodeficiency virus (HIV) (4-7).We have isolated (8) the cDNA and the gene encoding T4 and have determined the nucleotide sequence of the fulllength cDNA clone. To define the mechanisms of interaction of T4 with the surface of both antigen-presenting cells as well as with HIV, we have further analyzed the sequence, structure, and expression of the human and mouse § T4 genes. Human T4, as well the mouse homologue L3T4, exhibit a polyimmunoglobulin-like structure with four tandem variable-joining (VJ)-like domains. This polyimmunoglobulinlike structure of T4 is homologous to an increasingly large number of recognition molecules. Moreover, we observe that T4 expression is not restricted to T cells, suggesting that T4 plays a more general role in cell-cell interactions. MATERIALS AND METHODSAll materials and procedures have been described (7-9). RESULTS T4 Exhibits a Polyimmunoglobulin-like Structur...