Biomedical subjects

Matthew D Rand

Publications and source records attributed to Matthew D Rand.

4 recordsLinked to original sources

PubMed2006

Methylmercury induces activation of Notch signaling.

Methylmercury (MeHg) toxicity in humans manifests deficits in neurological function. Cases of prenatal exposure to mercury have established that the developing nervous system is most highly susceptible to perturbation by MeHg. At a cellular level, MeHg-induced defects result from altered neuronal proliferation, migration and pathfinding. However, the molecular targets of MeHg that give rise to these outcomes are not fully understood. In an overall effort to identify the fundamental molecular targets of MeHg in neural development, we have explored the effects of MeHg on cell surface receptor function using the simplified Drosophila model. In this study, we investigated the potential role of MeHg to alter activity of the Notch receptor pathway, a highly conserved cell-cell signaling mechanism that controls cell fate decisions, proliferation, migration and neurite outgrowth in neural development. Notch receptor activation requires proteolysis by a cell surface ADAM metalloprotease. ADAM proteases are required for normal neural development and are activated by organomercurials, thus presenting a possible mechanism for MeHg neurotoxicity. Here, we demonstrate a concentration- and time-dependent increase in Notch receptor activity with MeHg exposure in three distinct Drosophila cell lines. Ten micromolar MeHg results in a 4-5.5-fold increase in Notch signaling as measured by the upregulation of two enhancer of split (E(spl)) target genes. MeHg-induced Notch activity also correlates with receptor proteolysis. Targeted knockdown of Notch protein expression demonstrates that MeHg induced E(spl) activation specifically requires the Notch receptor. Furthermore, MeHg-induced Notch activity is partially attenuated by the metalloprotease inhibitor, GM6001, consistent with a model in which MeHg promotes activation of ADAM metalloproteases. Finally, we demonstrate that inorganic HgCl(2) is significantly less active in inducing Notch activity, suggesting a mechanism specific to organic species of mercury. Overall, these data identify Notch as a potential target for MeHg toxicity in the developing nervous system.

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PubMed2006

Endocytosis-independent mechanisms of Delta ligand proteolysis.

Delta proteins function as cell surface ligands for Notch receptors in a highly conserved signal transduction mechanism. Delta activates Notch by "trans-endocytosis", whereby endocytosis of Delta that is in complex with Notch on a neighboring cell induces activating cleavages in Notch. Alternatively, proteolysis of Delta renders the ligand inactive by dissociating the extracellular and cytosolic domains. How proteolysis and trans-endocytosis cooperate in Delta function is not well understood. We now show that Drosophila Delta proteolysis occurs independent of and prior to endocytosis in neuroblasts and ganglion mother cells in vivo and cells in culture. Delta cleavage occurs at two novel sites that we identify in the juxtamembrane (JM) and transmembrane (TM) domains. In addition to the previously identified Kuzbanian ADAM protease, which acts on the JM domain, proteolysis in the TM domain is facilitated by a thiol-sensitive aspartyl protease that is distinct from Presenilin. Furthermore, cleavage in the TM domain is upregulated in the presence of Notch. Overall, Drosophila Delta proteolysis differs from the conventional regulated intramembrane proteolysis (RIP) mechanism by two criteria: (1) TM-domain processing of Delta is not sensitive to Presenilin, and (2) TM and JM domain cleavages occur independently of each other. Altogether, these data support a model whereby proteolysis can modulate Delta ligand activity independently of endocytosis.

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PubMed2003

Notch-induced proteolysis and nuclear localization of the Delta ligand.

The Delta protein is a single-pass transmembrane ligand for the Notch family of receptors. Delta binding to Notch invokes regulated intramembrane proteolysis and nuclear translocation of the Notch intracellular domain. Delta is proteolytically processed at two sites, Ala(581) and Ala(593) in the juxtamembrane and transmembrane domains, respectively (Mishra-Gorur, K., Rand, M. D., Perez-Villamil, B., and Artavanis-Tsakonas, S. (2002) J. Cell Biol. 159, 313-324). Controversy over the role of Delta processing in propagating Notch signals has stemmed from conflicting reports on the activity or inactivity of soluble extracellular domain products of Delta. We have examined Delta proteolysis in greater detail and report that Delta undergoes three proteolytic cleavages in the region of the juxtamembrane and transmembrane domains. Only one of these cleavages, analogous to cleavage at Ala(581), is dependent on the Kuzbanian ADAM metalloprotease. The two additional cleavages correspond to the previously described cleavage at Ala(593) and a novel unidentified site within or close to the transmembrane domain. Delta processing is up-regulated in co-cultures with Notch-expressing cells and is similarly induced by p-aminophenylmercuric acetate, a well documented activator of metalloproteases. Furthermore, expression of a truncated intracellular isoform of Delta shows prominent nuclear localization. Altogether, these data demonstrate a role for Notch in inducing Delta proteolysis and implicate a nuclear function for Delta, consistent with a model of bi-directional signaling through Notch-Delta interactions.

Alanine↗

PubMed2002

Down-regulation of Delta by proteolytic processing.

Notch signaling regulates cell fate decisions during development through local cell interactions. Signaling is triggered by the interaction of the Notch receptor with its transmembrane ligands expressed on adjacent cells. Recent studies suggest that Delta is cleaved to release an extracellular fragment, DlEC, by a mechanism that involves the activity of the metalloprotease Kuzbanian; however, the functional significance of that cleavage remains controversial. Using independent functional assays in vitro and in vivo, we examined the biological activity of purified soluble Delta forms and conclude that Delta cleavage is an important down-regulating event in Notch signaling. The data support a model whereby Delta inactivation is essential for providing the critical ligand/receptor expression differential between neighboring cells in order to distinguish the signaling versus the receiving partner.

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