title: TPC1 has two variant isoforms, and their removal has different effects on endo-lysosomal functions compared to loss of TPC2.
abstract: Organelle ion homeostasis within the endo-lysosomal system is critical for physiological functions. Two-pore channels (TPCs) are cation channels that reside in endo-lysosomal organelles, and overexpression results in endo-lysosomal trafficking defects. However, the impact of a lack of TPC expression on endo-lysosomal trafficking is unknown. Here, we characterize Tpcn1 expression in two transgenic mouse lines (Tpcn1(XG716) and Tpcn1(T159)) and show expression of a novel evolutionarily conserved Tpcn1B transcript from an alternative promoter, raising important questions regarding the status of Tpcn1 expression in mice recently described to be Tpcn1 knockouts. We show that the transgenic Tpcn1(T159) line lacks expression of both Tpcn1 isoforms in all tissues analyzed. Using mouse embryonic fibroblasts (MEFs) from Tpcn1(-/-) and Tpcn2(-/-) animals, we show that a lack of Tpcn1 or Tpcn2 expression has no significant impact on resting endo-lysosomal pH or morphology. However, differential effects in endo-lysosomal function were observed upon the loss of Tpcn1 or Tpcn2 expression; thus, while Tpcn1(-/-) MEFs have impaired trafficking of cholera toxin from the plasma membrane to the Golgi apparatus, Tpcn2(-/-) MEFs show slower kinetics of ligand-induced platelet-derived growth factor receptor β (PDGFRβ) degradation, which is dependent on trafficking to lysosomes. Our findings indicate that TPC1 and TPC2 have important but distinct roles in the endo-lysosomal pathway.
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom margarida.ruas@pharm.ox.ac.uk john.parrington@pharm.ox.ac.uk.
authors.forename: Margarida
authors.initials: M
authors.lastname: Ruas
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Kai-Ting
authors.initials: KT
authors.lastname: Chuang
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Lianne C
authors.initials: LC
authors.lastname: Davis
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Areej
authors.initials: A
authors.lastname: Al-Douri
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Patricia W
authors.initials: PW
authors.lastname: Tynan
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Ruth
authors.initials: R
authors.lastname: Tunn
authors.affiliation: The Mary Lyon Centre, MRC Harwell, Harwell, United Kingdom.
authors.forename: Lydia
authors.initials: L
authors.lastname: Teboul
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom.
authors.forename: Antony
authors.initials: A
authors.lastname: Galione
authors.affiliation: Department of Pharmacology, University of Oxford, Oxford, United Kingdom margarida.ruas@pharm.ox.ac.uk john.parrington@pharm.ox.ac.uk.
authors.forename: John
authors.initials: J
authors.lastname: Parrington
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D001483:Base Sequence; D015220:Calcium Channels / Q000235:genetics* / Q000378:metabolism*; D002462:Cell Membrane / Q000502:physiology; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000378:metabolism; D017124:Conserved Sequence; D055786:Gene Knockout Techniques; D006056:Golgi Apparatus / Q000502:physiology; D008247:Lysosomes / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000378:metabolism; D000097903:Two-Pore Channels
chemical_list: D015220:Calcium Channels; D002772:Cholera Toxin; D020033:Protein Isoforms; D020797:Receptor, Platelet-Derived Growth Factor beta; C546784:TPCN1 protein, mouse; D000097903:Two-Pore Channels; C546785:TPCN2 protein, mouse