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Biomedical subjects

Rohan D Teasdale

Publications and source records attributed to Rohan D Teasdale.

12 recordsLinked to original sources
PubMed2003

Contextual binding of p120ctn to E-cadherin at the basolateral plasma membrane in polarized epithelia.

E-cadherin-catenin complexes mediate cell-cell adhesion on the basolateral membrane of epithelial cells. The cytoplasmic tail of E-cadherin supports multiple protein interactions, including binding of beta-catenin at the C terminus and of p120ctn to the juxtamembrane domain. The temporal assembly and polarized trafficking of the complex or its individual components to the basolateral membrane are not fully understood. In Madin-Darby canine kidney cells at steady state and after treatment with cycloheximide or temperature blocks, E-cadherin and beta-catenin localized to the Golgi complex, but p120ctn was found only at the basolateral plasma membrane. We previously identified a dileucine sorting motif (Leu586-Leu587, termed S1) in the juxtamembrane domain of E-cadherin and now show that it is required to target full-length E-cadherin to the basolateral membrane. Removal of S1 resulted in missorting of E-cadherin mutants (EcadDeltaS1) to the apical membrane; beta-catenin was simultaneously missorted and appeared at the apical membrane. p120ctn was not mistargeted with EcadDeltaS1, but could be recruited to the E-cadherin-catenin complex only at the basolateral membrane. These findings help define the temporal assembly and sorting of the E-cadherin-catenin complex and show that membrane recruitment of p120ctn in polarized cells is contextual and confined to the basolateral membrane.

Amino Acid Motifs↗
PubMed2003

Genomic screen for genes involved in mammalian craniofacial development.

Using a subtractive hybridisation approach, we enriched for genes likely to play a role in embryonic development of the mammalian face and other structures. This was achieved by subtracting cDNA derived from adult mouse liver from that derived from 10.5 dpc mouse embryonic branchial arches 1 and 2. Random sequencing of clones from the resultant library revealed that a high percentage correspond to genes with a previously established role in embryonic development and disease, while 15% represent novel or uncharacterised genes. Whole mount in situ hybridisation analysis of novel genes revealed that approximately 50% have restricted expression during embryonic development. In addition to expression in branchial arches, these genes showed a range of expression domains commonly including neural tube and somites. Notably, all genes analysed were found to be expressed not only in the branchial arches but also in the developing limb buds, providing support for the hypothesis that development of the limbs and face is likely to involve analogous molecular processes.

Animals↗
PubMed2003

Intracellular sorting and transport of proteins.

The secretory and endocytic pathways of eukaryotic organelles consist of multiple compartments, each with a unique set of proteins and lipids. Specific transport mechanisms are required to direct molecules to defined locations and to ensure that the identity, and hence function, of individual compartments are maintained. The localisation of proteins to specific membranes is complex and involves multiple interactions. The recent dramatic advances in understanding the molecular mechanisms of membrane transport has been due to the application of a multi-disciplinary approach, integrating membrane biology, genetics, imaging, protein and lipid biochemistry and structural biology. The aim of this review is to summarise the general principles of protein sorting in the secretory and endocytic pathways and to highlight the dynamic nature of these processes. The molecular mechanisms involved in this transport along the secretory and endocytic pathways are discussed along with the signals responsible for targeting proteins to different intracellular locations.

Animals↗
PubMed2003

Analysis of the mouse transcriptome for genes involved in the function of the nervous system.

We analyzed the mouse Representative Transcript and Protein Set for molecules involved in brain function. We found full-length cDNAs of many known brain genes and discovered new members of known brain gene families, including Family 3 G-protein coupled receptors, voltage-gated channels, and connexins. We also identified previously unknown candidates for secreted neuroactive molecules. The existence of a large number of unique brain ESTs suggests an additional molecular complexity that remains to be explored.A list of genes containing CAG stretches in the coding region represents a first step in the potential identification of candidates for hereditary neurological disorders.

Adenine↗
PubMed2003

Mouse proteome analysis.

A general overview of the protein sequence set for the mouse transcriptome produced during the FANTOM2 sequencing project is presented here. We applied different algorithms to characterize protein sequences derived from a nonredundant representative protein set (RPS) and a variant protein set (VPS) of the mouse transcriptome. The functional characterization and assignment of Gene Ontology terms was done by analysis of the proteome using InterPro. The Superfamily database analyses gave a detailed structural classification according to SCOP and provide additional evidence for the functional characterization of the proteome data. The MDS database analysis revealed new domains which are not presented in existing protein domain databases. Thus the transcriptome gives us a unique source of data for the detection of new functional groups. The data obtained for the RPS and VPS sets facilitated the comparison of different patterns of protein expression. A comparison of other existing mouse and human protein sequence sets (e.g., the International Protein Index) demonstrates the common patterns in mammalian proteomes. The analysis of the membrane organization within the transcriptome of multiple eukaryotes provides valuable statistics about the distribution of secretory and transmembrane proteins

Animals↗
PubMed2003

The mouse secretome: functional classification of the proteins secreted into the extracellular environment.

We have developed a computational strategy to identify the set of soluble proteins secreted into the extracellular environment of a cell. Within the protein sequences predominantly derived from the RIKEN representative transcript and protein set, we identified 2033 unique soluble proteins that are potentially secreted from the cell. These proteins contain a signal peptide required for entry into the secretory pathway and lack any transmembrane domains or intracellular localization signals. This class of proteins, which we have termed the mouse secretome, included >500 novel proteins and 92 proteins <100 amino acids in length. Functional analysis of the secretome included identification of human orthologs, functional units based on InterPro and SCOP Superfamily predictions, and expression of the proteins within the RIKEN READ microarray database. To highlight the utility of this information, we discuss the CUB domain-containing protein family.

Animals↗
PubMed2002

The phox homology (PX) domain-dependent, 3-phosphoinositide-mediated association of sorting nexin-1 with an early sorting endosomal compartment is required for its ability to regulate epidermal growth factor receptor degradation.

Recent studies have shown that phox homology (PX) domains act as phosphoinositide-binding motifs. The majority of PX domains studied show binding to phosphatidylinositol 3-monophosphate (PtdIns(3)P), an association that allows the host protein to localize to membranes of the endocytic pathway. One issue, however, is whether PX domains may have alternative phosphoinositide binding specificities that could target their host protein to distinct subcellular compartments or allow their allosteric regulation by phosphoinositides other than PtdIns(3)P. It has been reported that the PX domain of sorting nexin 1 (SNX1) specifically binds phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) (Zhong, Q., Lazar, C. S., Tronchere, H., Sato, T., Meerloo, T., Yeo, M., Songyang, Z., Emr, S. D., and Gill, G. N. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 6767-6772). In the present study, we have shown that whereas SNX1 binds PtdIns(3,4,5)P(3) in protein:lipid overlay assays, in liposomes-based assays, binding is observed to PtdIns(3)P and phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P(2)) but not to PtdIns(3,4,5)P(3). To address the significance of PtdIns(3,4,5)P(3) binding, we examined the subcellular localization of SNX1 under conditions in which plasma membrane PtdIns(3,4,5)P(3) levels were significantly elevated. Under these conditions, we failed to observe association of SNX1 with this membrane. However, consistent with the binding to PtdIns(3)P and PtdIns(3,5)P(2) being of more physiological significance was the observation that the association of SNX1 with an early endosomal compartment was dependent on a 3-phosphoinositide-binding PX domain and the presence of PtdIns(3)P on this compartment. Finally, we have shown that the PX domain-dependent/early endosomal association of SNX1 is important for its ability to regulate the targeting of internalized epidermal growth factor receptor for lysosomal degradation.

Animals↗
PubMed2002

Targeting of the GRIP domain to the trans-Golgi network is conserved from protists to animals.

The GRIP domain, found in a family of coiled-coil peripheral membrane Golgi proteins, is a specific targeting sequence for the trans-Golgi network of animal cells. In this study we show that a coiled-coil protein with a GRIP domain occurs in the primitive eukaryote, Trypanosoma brucei, and that reporter proteins containing this domain can be used as a marker for the poorly characterized trans Golgi/trans-Golgi network of trypanosomatid parasites. The T. brucei GRIP domain, when fused to the carboxyl terminus of the green fluorescent protein (GFP-TbGRIP), was efficiently localized to the Golgi apparatus of transfected COS cells. Overexpression of GFP-TbGRIP in COS cells displaced the endogenous GRIP protein, GCC1p, from the Golgi apparatus indicating that the trypanosomatid and mammalian GRIP sequences interact with similar membrane determinants. GFP fusion proteins containing either the T. brucei GRIP domain or the human p230 GRIP (p230GRIP) domain were also expressed in the trypanosomatid parasite, Leishmania mexicana, and localized by fluorescence and immuno-electron microscopy to the trans face of the single Golgi apparatus and a short tubule that extended from the Golgi apparatus. Binding of GFP-p230GRIP to Golgi membranes in L. mexicana was abrogated by mutation of a critical tyrosine residue in the p230 GRIP domain. The levels of GFP-GRIP fusion proteins were dramatically reduced in stationary-phase L. mexicana promastigotes, suggesting that specific Golgi trafficking steps may be down-regulated as the promastigotes cease dividing. This study provides a protein marker for the trans-Golgi network of trypanosomatid parasites and suggests that the GRIP domain binds to a membrane component that has been highly conserved in eukaryotic evolution.

Amino Acid Sequence↗
PubMed2002

Prediction of Golgi Type II membrane proteins based on their transmembrane domains.

MOTIVATION: A major issue in cell biology today is how distinct intracellular regions of the cell, like the Golgi Apparatus, maintain their unique composition of proteins and lipids. The cell differentially separates Golgi resident proteins from proteins that move through the organelle to other subcellular destinations. We set out to determine if we could distinguish these two types of transmembrane proteins using computational approaches. RESULTS: A new method has been developed to predict Golgi membrane proteins based on their transmembrane domains. To establish the prediction procedure, we took the hydrophobicity values and frequencies of different residues within the transmembrane domains into consideration. A simple linear discriminant function was developed with a small number of parameters derived from a dataset of Type II transmembrane proteins of known localization. This can discriminate between proteins destined for Golgi apparatus or other locations (post-Golgi) with a success rate of 89.3% or 85.2%, respectively on our redundancy-reduced data sets. AVAILABILITY: See http://microarray.imb.uq.edu.au/golgi/

Algorithms↗
PubMed2002

Secretory pathway of trypanosomatid parasites.

The Trypanosomatidae comprise a large group of parasitic protozoa, some of which cause important diseases in humans. These include Trypanosoma brucei (the causative agent of African sleeping sickness and nagana in cattle), Trypanosoma cruzi (the causative agent of Chagas' disease in Central and South America), and Leishmania spp. (the causative agent of visceral and [muco]cutaneous leishmaniasis throughout the tropics and subtropics). The cell surfaces of these parasites are covered in complex protein- or carbohydrate-rich coats that are required for parasite survival and infectivity in their respective insect vectors and mammalian hosts. These molecules are assembled in the secretory pathway. Recent advances in the genetic manipulation of these parasites as well as progress with the parasite genome projects has greatly advanced our understanding of processes that underlie secretory transport in trypanosomatids. This article provides an overview of the organization of the trypanosomatid secretory pathway and connections that exist with endocytic organelles and multiple lytic and storage vacuoles. A number of the molecular components that are required for vesicular transport have been identified, as have some of the sorting signals that direct proteins to the cell surface or organelles in the endosome-vacuole system. Finally, the subcellular organization of the major glycosylation pathways in these parasites is reviewed. Studies on these highly divergent eukaryotes provide important insights into the molecular processes underlying secretory transport that arose very early in eukaryotic evolution. They also reveal unusual or novel aspects of secretory transport and protein glycosylation that may be exploited in developing new antiparasite drugs.

Animals↗
PubMed2002

Conserved modularity and potential for alternate splicing in mouse and human Slit genes.

The vertebrate Slit gene family currently consists of three members; Slit1, Slit2 and Slit3. Each gene encodes a protein containing multiple epidermal growth factor and leucine rich repeat motifs, which are likely to have importance in cell-cell interactions. In this study, we sought to fully define and characterise the vertebrate Slit gene family. Using long distance PCR coupled with in silico mapping, we determined the genomic structure of all three Slit genes in mouse and man. Analysis of EST and genomic databases revealed no evidence of further Slit family members in either organism. All three Slit genes were encoded by 36 (Slit3) or 37 (Slit1 and Slit2) exons covering at least 143 kb or 183 kb of mouse or human genomic DNA respectively. Two additional potential leucine-rich repeat encoding exons were identified within intron 12 of Slit2. These could be inserted in frame, suggesting that alternate splicing may occur in Slit2. A search for STS sequences within human Slit3 anchored this gene to D5S2075 at the 5' end (exon 4) and SGC32449 within the 3' UTR, suggesting that Slit3 may cover greater than 693 kb. The genomic structure of all Slit genes demonstrated considerable modularity in the placement of exon-intron boundaries such that individual leucine-rich repeat motifs were encoded by individual 72 bp exons. This further implies the potential generation of multiple Slit protein isoforms varying in their number of repeat units. cDNA library screening and EST database searching verified that such alternate splicing does occur.

3' Untranslated Regions↗