Connectin

EVOLUTIONARY HOMOLOGS

Drosophila LRR proteins

The gene tlr (for Toll-like receptor) that encodes a protein containing multiple LRRs in its presumed extracellular domain, has a single transmembrane segment and homology to the cytoplasmic domain of the interleukin 1 receptor in its presumed intracellular domain. The pattern of tlr expression at the extended germ band stage is characterized by 15 transverse stripes in the gnathal and trunk segments, with four patches of expression corresponding to head segments and an additional patch of expression in the presumptive hindgut. The segmentally repeated tlr stripes in the trunk overlap both the wingless and engrailed stripes and thus span the parasegment boundary. The tlr stripes require pair rule gene function for their establishment and later become dependent upon segment-polarity gene function for their maintenance. Later segmental modulation of tlr expression later in the tracheal system is dependent upon the function of the homeotic genes of the bithorax complex. tlris also prominently expressed in imaginal discs. In the eye disc, this expression occurs in two stripes at the anterior and posterior margins of the morphogenetic furrow; this expression is consistent with a genetic interaction between a tlr mutation and an eye-specific allele of hedgehog. All of these data combine to suggest a role for tlr in interactions between cells at critical boundaries during development (Chiang, 1994).

Two members of a novel class of genes have been identified in Drosophila that encode putative transmembrane proteins with six leucine-rich repeats and a single immunoglobulin loop. These two molecules, Kekkon1 and Kekkon2, show striking conservation in their extracellular domains and have large and more divergent intracellular regions. Both genes are expressed in neurons as they differentiate in the embryonic central nervous system (CNS). kek1 is also expressed in other patterned epithelia, such as the follicle cells of the developing egg chamber, where it is found in a dorsal-ventral gradient around the oocyte. The homology of the two kek genes to other known adhesion and signaling molecules, together with their expression patterns, suggests that both genes are involved in interactions at the cell surface. Genetic analysis reveals that deletion of the kek1 gene causes no obvious developmental defects. The coexpression of kek2 in the CNS suggests that Kek1 is part of a family of cell surface proteins with redundant function (Musacchio, 1996).

The Drosophila tartan gene is transcribed in an unusual embryonic pattern of intersecting stripes that are generated in response to the anterior-posterior and dorsal-ventral regulatory systems. tartan encodes a putative transmembrane protein containing extracellular leucine-rich repeats characteristic of numerous cell surface receptors and adhesion proteins. Its expression is correlated with aspects of segmentation and neurogenesis, including the formation of neuroblasts, sensory mother cells, and peripheral nerves. Mutants homozygous for a recessive lethal tartan loss-of-function allele exhibit defects in the position and number of cells within peripheral sense organs, the routing of peripheral nerves, and the organization of commissures within the central nervous system. Mutants are also defective in muscle organization. These results suggest that tartan is required for cell surface interactions important for normal organization of epidermal and subepidermal structures (Chang, 1993).

The sequence of a Drosophila embryonic LRR protein (LRR47) includes a hydrophobic N-terminal that may constitute an ER signal sequence, eight LRR copies and a unique C-terminal. The transcript of the LRR47 gene is detected in adult females and in early embryogenesis. It is not found in adult males and is only present at low levels in embryos after 6 h of development. In Western blot experiments, a protein of approx. 47 kDa, which is expressed in a similar developmental profile and purifies in peripheral membrane protein extracts, is detected by an antibody specific for LRR47. The LRR47 gene maps to position 32A on the left arm of chromosome 2, an interval in which three genes with semi-lethal maternal effects (dal, hup and wdl) are located (Ntwasa, 1994).

Upon reaching the target region, neuronal growth cones transiently search through them and form synaptic connections with only a subset of these potential targets. The capricious (caps) gene may regulate these processes in Drosophila. caps encodes a transmembrane protein with 14 leucine-rich repeats (LRRs) in its extracellular domain. Among the proteins with LRRs, Caps protein is most closely related to the product of the Drosophila gene tartan, with amino acid similarity extending beyond the LRR region into the cytoplasmic region. Another LRR protein implicated in Drosophila neuromuscular recognition is Connectin, which is expressed both pre- and post-synaptically on a subset of motorneurons and muscles different from those involved in Capricious or Tartan expression. Capricious LRRs are 67% homologous to those of Tartan, which contains a longer intracellular domain, but only 20% homologous to those of Connectin, which is attached to the cell membrane by a GPI anchor. During the formation of neuromuscular synapses, caps is expressed in a small number of synaptic partners, including muscle 12 and the motorneurons that innervate it. Loss-of-function and ectopic expression of caps alter the target specificity of muscle 12 motorneurons, indicating a role for caps in selective synapse formation. In wild-type larvae, muscle 12 is innervated by the terminal branch of ISNb, including the RP5 axon, which projects to the boundary between muscles 12 and 13 and forms synaptic endings exclusively on muscle 12. In contrast, in caps mutant larvae, the terminal branch is often accompanied by additional varicosities on muscle 13, a neighboring caps-negative muscle. Ectopic overexpression of caps causes formation of more ectopic synapses. The ISNb terminal forms one or more additional collaterals that form more robust synaptic endings on muscle 13. The ectopic nerve endings contain type III boutons, which are typical of muscle 12 but not muscle 13 neuromuscular synapses. The expression of caps on both sides of the synaptic partners suggests that caps functions homophilically, as has been proposed for the candidate target recognition molecules Connectin and Fasciclin III. However, expression of caps in S2 cells does not promote cell aggregation (Shishido, 1998).

Cell surface LRR proteins

LIG-1 is up-regulated during neural differentiation in mouse P19 embryonal carcinoma cells. Comparative sequence analysis reveals LIG-1 to be a novel integral membrane glycoprotein (1091 amino acids) containing an extracellular region (794 amino acids) with a potential signal peptide, 15 leucine-rich repeats, 3 immnunoglobulin-like domains, and 7 potential N-glycosylation sites, a transmembrane region of 23 amino acids, and a cytoplasmic region of 274 amino acids. This protein, therefore, is a new member of both the leucine-rich repeat and the immunoglobulin superfamilies. Furthermore, Northern blot and in situ hybridization analyses showed LIG-1 gene expression to be predominantly in the brain, restricted to a small subset of glial cells such as Bergmann glial cells of the cerebellum and glial cells in the nerve fiber layer of the olfactory bulb. On the basis of its structural features and expression pattern, it is proposed that LIG-1 functions as a cell type-specific adhesion molecule or receptor at the glial cell surface, and plays a role in several aspects of nervous system development, including neuroglial differentiation and the development and/or maintenance of neural functions where it is expressed (Suzuki, 1996).

A neonatal mouse brain cDNA library was screened with a human partial cDNA encoding LRR protein as a probe. Two independent cDNAs were obtained encoding LRR proteins, designated NLRR-1 and NLRR-2 (Neuronal Leucine-Rich Repeat proteins). These two clones are about 60% homologous to one another, and NLRR-1 protein is a transmembrane protein. Both NLRR-1 and NLRR-2 mRNAs are expressed primarily in the central nervous system (CNS); NLRR-1 mRNA is also detected in non-neuronal tissues such as cartilage, while NLRR-2 mRNA expression is confined to the CNS at all developmental stages. These results suggest that there is at least one LRR protein family in the mouse and that these molecules may play significant but distinct roles in neural development and in the adult nervous system (Taguchi, 1996).

Using a human brain cDNA fragment encoding a LRR as a probe, a mouse brain cDNA was isolated that encodes a new LRR protein: NLRR-3 protein. The isolated cDNA is 3350 bp long including one open reading frame encoding a protein of 707 amino acids, the deduced amino acid sequence of which has a signal peptide and a transmembrane region. The NLRR-3 protein also contains an RGD sequence and 11 LRRs with amino- and carboxy-terminal LRR-flanking regions that are conserved among adhesive proteins and signal-transducing receptors in this family. Northern-blot analysis reveals strong expression of an approx. 4.2 kb NLRR-3 mRNA in the brain from E17 to P7, and weak expression in adults. NLRR-3 mRNA is expressed in the brain, in which stronger expression is localized in the ventricular zone and anlage of thalamus, spinal cord, and dorsal root ganglion in E11-17 cerebellum, and the cerebral cortex in adults. The molecular structure, in addition to the transient and localized expression, suggests that the NLRR-3 protein plays a role in the development and maintenance of the nervous system by protein-protein interactions (Taniguchi, 1996).

A new human gene, named GARP, is localized in the 11q14 chromosomal region. GARP, comprising two coding exons, is expressed as two major transcripts of 4.4 and 2.8 kilobases, respectively, and encodes a putative transmembrane protein of 662 amino acids, the extracellular portion of which is almost entirely made of leucine-rich repeats. The molecular weight of the protein immunoprecipitated from transfected cells is 80,000. The GARP protein has structural similarities to the human GP Ib alpha and GP V platelet proteins, and with Drosophila adhesion molecules of the Chaoptin, Toll, and Connectin proteins (Ollendorff, 1994).

The organization and pattern of expression of the mouse Garp gene is composed of two coding exons, expressed as a major 4.3 kb mRNA, encoding a putative LRR transmembrane protein with an extracellular region almost entirely made of 20 repeats, and a short intracytoplasmic region. The mouse GARP deduced amino-acid sequence is highly similar to that of the human protein. The Garp gene is expressed in various areas in the mid-gestation developing embryo, including skin, lens fiber cells, nasal cavity, smooth and skeletal muscles, lung, and megakaryocytes of the fetal liver. In the adult it is expressed in the megakaryocytes of the spleen and in endothelial cells of the placenta. The data suggests that GARP might be involved in platelet-endothelium interactions (Roubin, 1996).

The RP105 Ag is a murine B cell surface molecule that transmits an activation signal into B cells following ligation with anti-RP105 mAb. The activation leads to protection of B cells from irradiation- or dexamethasone-induced apoptosis, and to B cell proliferation. A cDNA encoding the RP105 Ag has been isolated. This protein is a type I transmembrane protein consisting of 641 amino acids in a mature form. The transcript is observed in spleen, but not in thymus, kidney, muscle, heart, brain, or liver. Stable transfection of the cDNA clone confers the expression of the RP105 Ag on a pro-B cell line. The RP105 molecule possesses 22 tandem repeats of a leucine-rich motif. Amino- and carboxyl-flanking regions that are characteristically conserved among members of the family are located on both sides of tandemly repeated leucine-rich motifs in RP105 molecule. These results demonstrate that RP105 is a novel member of the leucine-rich repeat protein family, and the first member that is specifically expressed on B cells (Miyake, 1995).

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