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Biomedical subjects

Suzanne M Watt

Publications and source records attributed to Suzanne M Watt.

4 recordsLinked to original sources
PubMed2003

Novel mesenchymal and haematopoietic cell isoforms of the SHP-2 docking receptor, PZR: identification, molecular cloning and effects on cell migration.

SHP-2 (Src homology phosphatase type-2) is essential for haematopoietic skeletal and vascular development. Thus the identification of its binding partners is critically important. In the present study, we describe a unique monoclonal antibody, WM78, which interacts with PZR, a SHP-2 binding partner. Furthermore, we identify two novel isoforms of PZR, PZRa and PZRb, derived by differential splicing from a single gene transcription unit on human chromosome 1q24. All are type 1 transmembrane glycoproteins with identical extracellular and transmembrane domains, but differ in their cytoplasmic tails. The PZR intracellular domain contains two SHP-2 binding immunoreceptor tyrosine-based inhibitory motifs (VIY(246)AQL and VVY(263)ADI) which are not present in PZRa and PZRb. Using the WM78 monoclonal antibody, which recognizes the common extracellular domain of the PZR isoforms, we demonstrate that the PZR molecules are expressed on mesenchymal and haematopoietic cells, being present on the majority of CD34(+)CD38(+) and early clonogenic progenitors, and at lower levels on CD34(+)CD38(-) cells and the hierarchically more primitive pre-colony forming units. Interestingly, we show by reverse transcriptase-PCR that the PZR isoforms are differentially expressed in haematopoietic, endothelial and mesenchymal cells. Both PZR and PZRb are present in CD133(+) precursors and endothelial cells, PZRb predominates in mesenchymal and committed myelomonocytic progenitor cells, and all three isoforms occur in erythroid precursor cell lines. Importantly, using SHP-2 mutant (Delta 46-110) and SHP-2 rescue of embryonic fibroblasts stably expressing the PZR isoforms, we demonstrate for the first time that PZR, but not PZRa or PZRb, facilitates fibronectin- dependent migration of cells expressing a competent SHP-2 molecule. These observations will be instrumental in determining the mechanisms whereby PZR isoforms regulate cell motility.

Amino Acid Motifs↗
PubMed2003

Colocalization analysis of sialomucins CD34 and CD164.

Flow cytometric protocols are employed to identify and characterize hemopoietic stem/progenitor populations before transplantation. Cell surface antigens, including CD34, are employed in this process and widely used in harvest protocols, which largely ignores the potential functional role of such antigens. Transmembrane glycoprotein sialomucins, including CD34 and CD164, have been implicated in cell-to-cell interactions and activation. CD164, also expressed on early hemopoietic populations, was reported to have a possible function facilitating CD34(+) cells to adhere to bone marrow stroma. In this study, we employed high-definition laser-scanning confocal microscopy to investigate CD34 and CD164 surface co-localization patterns on bone marrow and cord blood cells and to compare the expression patterns using a three-dimensional computer-generated method developed in house. Differential interference microscopy analysis revealed bone marrow membrane activity was higher than the corresponding cord blood counterpart, perhaps indicating the marrow microenvironmental nature. Fluorescence analysis of CD34 and CD164 antigens showed both were expressed first in a halo-like pattern and second in antigen-dense pockets. Three-dimensional computer analyses further revealed that this pocketing corresponded to dense crest-like surface structures appearing to rise from the point of adherence on the slide. Further, it was found that CD34 and CD164 display strong colocalization patterns on cells expressing both antigens. The dual nature of the CD34 and CD164 antigens discovered here lends further evidence to the previous literature implicating a strong functional link between these two sialomucins, which should be considered in the transplantation arena and in the function of such sialomucins as negative regulators of cell proliferation.

Antigens, CD34↗
PubMed2002

Human Siglec-5: tissue distribution, novel isoforms and domain specificities for sialic acid-dependent ligand interactions.

Human Siglec-5 is a sialic acid binding immunoglobulin (Ig)-like lectin (Siglec), comprising one N-terminal IgV-SET domain followed by three IgC2-SET domains, and a cytoplasmic domain with ITIM and SAP motifs which regulate cell signalling. We report the differential distribution of hSiglec-5 on neutrophil and macrophage subsets in tissues using monoclonal antibodies, 1A5 and 2H8, which require the first IgC2-SET domain for binding. Interestingly, hSiglec-5 was especially prominent on macrophages in reactive lymph nodes. We have identified four isoforms of hSiglec-5 possessing three (hSiglec-5-3L and -3C) or four (hSiglec-5-4L and -4S) extracellular domains linked to long (hSiglec-5-3L and -4L) or short (hSiglec-5-4S) cytoplasmic tails or existing as a soluble isoform (hSiglec-5-3C). hSiglec-5-4L has the broadest tissue distribution, being detected in adult spleen, thymus, lymph node, peripheral blood leucocytes and bone marrow, and in fetal lung and liver. A soluble Fc chimaeric protein containing the hSiglec-5-4L extracellular domain binds in a sialic acid-dependent manner to glycophorin A on human erythrocytes and to alpha2-3- and alpha2-6-sialyllactose moieties. Domain deletion mutants of hSiglec-5(D1-4)-Fc reveal that the first three IgC2-SET domains are required for optimal binding, with adhesion being abolished if the first IgC2-SET domain is deleted. This indicates that each hSiglec-5 isoform will interact with sialic acid ligands and provides the first step towards defining structure-function relationships of hSiglec-5 isoforms.

Amino Acid Sequence↗