Expression and Characterization of Human Transient Receptor Potential Melastatin 3 (hTRPM3)

Abstract

Transient receptor potential (TRP) cation-selective channels are an emerging class of proteins that are involved in a variety of important biological functions including pain transduction, thermosensation, mechanoregulation, and vasorelaxation. Utilizing a bioinformatics approach, we have identified the full-length human TRPM3 (hTRPM3) as a member of the TRP family. Following the identification of the founding member of this family, dTRP, which is from a Drosophila mutant with abnormal visual signal transduction (2), mammalian homologues have been cloned and all of them contain a six-transmembrane domain followed by a TRP motif (XWKFXR). Based on homology, they are divided into three subfamilies: TRPC (canonical), TRPV (vanilloid), and TRPM (melastatin) (3). Members of the TRPM subfamily have unusually long cytoplasmic tails at both ends of the channel domain, and some of the family members have an enzyme domain in the C-terminal region. Despite their similarities of structure, TRPMs have different ion-conductive properties, activation mechanisms, and putative biological functions. TRPM1 is down-regulated in metastatic melanomas (4). TRPM2 is a Ca 2ϩ -permeable channel that contains an ADP-ribose pyrophosphatase domain and can be activated by ADP-ribose, NAD (5, 6), and changes in redox status (7). The TRPM2 gene is mapped to the chromosome region linked to bipolar affective disorder, nonsyndromic hereditary deafness, Knobloch syndrome, and holosencephaly (8). Two splice variants of TRPM4 have been described. TRPM4a is predominantly a Ca 2ϩ -permeable channel (9); whereas TRPM4b conducts monovalent cations upon activation by changes in intracellular Ca 2ϩ (10). TRPM5 is associated with Beckwith-Wiedemann syndrome and a predisposition to neoplasias (11). TRPM7, another bifunctional protein, has kinase activity in addition to its ion channel activity. TRPM7 is regulated by Mg 2ϩ -ATP and/or inositol 1,4,5-disphosphate and is required for cell viability (12-14). TRPM8 is up-regulated in prostate cancer and other malignancies (15). Recently, it has been shown to be a receptor that senses cold stimuli (16,17).Using a bioinformatics approach, we have identified a member of the human TRPM subfamily that we have called hTRPM3, consistent with the unified TRP nomenclature (3). hTRPM3 contains long N and C termini, although it does not contain any additional enzymatic features. hTRPM3 mRNA is expressed primarily in kidney with lower levels in brain, testis, and spinal cord. When expressed in HEK 293 cells, hTRPM3 is co-localized with the plasma membrane and is capable of mediating Ca 2ϩ entry. This hTRPM3-mediated Ca 2ϩ conductance * The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.The nucleotide sequence (s)

Ca2+-independent Phospholipase A2-dependent Gating of TRPM8 by Lysophospholipids

“…Our single channel analysis showing differential effects on varying channel conformational states adds to the recent significant developments concerning the requirement of different TRPM8 residues for activation by cold, PIP 2 , and menthol (6,26). It should be noted that store-dependent mechanisms have recently been shown to contribute to the activation of such "apparently store-independent" TRP members as TRPM3 (27) and noxious cold-sensitive ANKTM1 (28), although the mediators in this mode of activation remain unknown.…”

Calmodulin and S100A1 Protein Interact with N Terminus of TRPM3 Channel

“…The TRPM3 ion channel has been proposed to play an important role in Ca 2ϩ homeostasis serving as a Ca 2ϩ entry pathway (2,5). To date, no regulation via Ca 2ϩ -binding proteins have been reported for this channel.…”

“…It is closely related to the first known member of the TRPM subfamily, TRPM1 (1). TRPM3 shares typical structural features with other TRP family members, such as six putative transmembrane-spanning domains, intracellular N and C termini, and a conserved TRP box and coiled-coil region in its C terminus (2,3). There are no ankyrin repeat domains present on its N-tail; instead, there is a so-called TRPM homology region, a roughly 700-amino acid-long domain only found in TRPM family members (4).…”

“…The channel is proposed to mediate a Ca 2ϩ concentrationdependent Ca 2ϩ entry pathway in kidney cells and thus is thought to play an important role in renal Ca 2ϩ homeostasis (2,5). Recently, TRPM3 was also reported to serve as a nociceptor channel involved in the detection of noxious heat (6).…”

The Transient Receptor Potential Superfamily of Ion Channels

“…TRPM3 is a Ca 2ϩ -permeable, nonselective cation channel whose activity is increased by hypotonicity (28). TRPM4 and TRPM5 are the only TRP channels that are permeable to monovalent cations but not Ca 2ϩ (P Ca /P Na Ͻ0.05:1) (29,30).…”