Eukaryotic cells have various intracellular organelles with distinct functions that are segregated by organellar membranes. Since organellar membranes, like the plasma membrane, are impermeable to ions, specialized membrane proteins such as ion transporters, pumps and channels play vital roles in the ion homeostasis of intra-organellar spaces as well as the cytosol.
Some proteins are targeted to multiple organelles before being delivered to their final destinations. For example, certain proteins, after being synthesized, travel from the ER (endoplasmic reticulum) to the Golgi apparatus, trans-Golgi network, secretory vesicles and eventually to the plasma membrane (secretory pathway). Furthermore, some proteins in the plasma membrane are internalized to specialized vesicles, called endosomes, before reaching their final location (endocytic pathway). The organellar pH along the secretory and endocytic pathways is tightly regulated, which is important in protein targeting, receptor-ligand interaction, and enzymatic activities in the organelle. It has been also suggested that impaired organellar pH may lead to cell death as well as unregulated proliferation and differentiation. However, the precise molecular mechanism underlying how organellar pH homeostasis is achieved is not yet understood.
My research interest concerns how organellar pH is regulated under physiological and pathological conditions. The Na+/H+ exchangers (NHEs) play pivotal roles in intracellular pH and cell volume regulation by their transporter function, exchanging extracellular Na+ for intracellular H+. Originally, the NHEs were identified in the plasma membrane as regulators of cytosolic pH. Recently, I have isolated human cDNAs encoding novel NHE isoforms localizing to organellar membranes that are involved in maintaining organellar pH and ion homeostasis. We will further characterize them by organellar ion flux assay and pH measurement in the presence of different pharmacological inhibitors. In addition to these cell-based techniques, in vitro reconstitution systems in liposomes will be developed for more precise functional analyses. We will also investigate NHE-interacting proteins through biochemical, cell biological, genetic and immunological techniques. To further define biological roles of NHE proteins as well as the interacting proteins, we are going to employ genetic approaches in different model-organism systems.
Zheng JC, Tham CT, Keatings K, Fan S, Liou AYC, Allan D, and Numata, M, “Secretory Carrier Membrane Protein (SCAMP) Deficiency Influences Behavior of Adult Flies.” Frontiers in Cell and Developmental Biology (In press)
Diering GH and Numata M, “Endosomal pH in Neuronal Signaling and Synaptic Transmission: Role of Na+/H+ Exchanger NHE5”, Frontiers in Membrane Physiology and Membrane Biophysics. 4 412 (2014)
Diering GH, Numata Y, Fan S, Church J and Numata M, “Endosomal Acidification by Na+/H+ Exchanger NHE5 Regulates TrkA Cell-Surface Targeting and NGF-Induced P13K Signaling.” Molecular Biology of the Cell, 24, 3435-48 (2013)
Onishi I, Lin PJ, Numata Y, Austin P, Cipollone J, Roberge M, Roskelley CD, Numata M. Organellar (Na+, K+)/H+ exchanger NHE7 regulates cell adhesion, invasion and anchorage-independent growth of breast cancer MDA-MB-231 cells. Oncol Rep. 2012 Feb;27(2):311-7.
Fonseca BD, Diering GH, Bidinosti MA, Dalal K, Alain T, Balgi AD, Forestieri, R. Nodwell M, Rajadurai CV, Gunaratnam C, Tee AR, Duong F, Andersen RJ, Orlowski J, Numata M, Sonenberg N, Roberge M. Structure-activity analysis of niclosamide reveals potential role for cytoplasmic pH in control of mammalian target of rapamycin complex 1 (mTORC1) signaling. J Biol Chem. 2012 May 18;287(21):17530-45.
Diering GH, Mills F, Bamji SX*, Numata M*. Regulation of dendritic spine growth through activity-dependent recruitment of the brain-enriched Na+/H+ exchanger NHE5. Mol Biol Cell. 2011 Jul 1;22(13):2246-57.
Balgi AD, Diering GH, Donohue E, Lam KK, Fonseca BD, Zimmerman C, Numata M, Roberge M. Regulation of mTORC1 signaling by pH. PLoS One. 2011;6(6):e21549
Sin WC, Moniz DM, Ozog MA, Tyler JE, Numata M, Church J. Regulation of early neurite morphogenesis by the Na+/H+ exchanger NHE1. J Neurosci. 2009 Jul 15;29(28):8946-59.
Diering GH, Church J, Numata M. Secretory Carrier Membrane Protein 2 Regulates Cell-surface Targeting of Brain-enriched Na+/H+ Exchanger NHE5. J Biol Chem. 2009 May 15;284(20):13892-903.
Kagami T, Chen S, Memar P, Choi M, Foster LJ, Numata M. Identification and biochemical characterization of the SLC9A7 interactome. Mol Membr Biol. 2008 Aug;25(5):436-47.
Onishi I, Lin PJ, Diering GH, Williams WP, Numata M. RACK1 associates with NHE5 in focal adhesions and positively regulates the transporter activity. Cell Signal. 2007 Jan;19(1):194-203.
Lin PJ, Williams WP, Kobiljski J, Numata M. Caveolins bind to (Na+, K+)/H+ exchanger NHE7 by a novel binding module. Cell Signal. 2007 May;19(5):978-988. (2007)
Onishi, I, Lin, PJC, Diering, GH, Williams, WP, and Numata, M. RACK1 associates with NHE5 and positively regulates the transporter activity Cell Signal 19 194-203 (2007)
Lin, PJC, Williams, WP, Luu, Y, Molday, RS, Orlowski, J, and Numata, M. Secretory Carrier Membrane Proteins interact and regulate the trafficking of the organellar Na+ /H+ exchanger NHE. J. Cell Sci 118 1885-97 (2005)
Szabo, EZ*, Numata, M*, Lukashova, V*, Iannuzzi, P, Orlowski,J. b -Arrestins bind and decrease cell-surface abundance of the Na+ /H+ exchanger NHE5 isoform J. Proc. Natl. Acad. Sci. USA 102 2790-2795 (2005)
Szaszi, K, Paulsen, A, Szabo, EZ, Numata, M, Grinstein, S and Orlowski, J. Clathrin-mediated endocytosis and recycling of the neural-specific Na+ /H+ exchanger NHE5 isoform: regulation by phosphatidylinositol 3′-kinase and the actin cytoskeleton J. Biol. Chem. 277 42623-42632 (2002)
Numata, M and Orlowski, J. Molecular cloning and characterization of a novel (Na+, K+ )/H+ exchanger localized to the trans -Golgi network J. Biol. Chem. 276 17387-17394 (2001)