McGill BIOL200 Nucleo-Cytoplasmic Transport Fall 2023 PDF

McGill BIOL200 - Fall 2023 Nucleo-Cytoplasmic Transport © R. Roy, 2023 McGill BIOL200 - Fall 2023 Nuclear pore complexes resemble baskets, and are the sites of nucleocytoplasmic transport Cytoplasmic face Nucleoplasmic face © R. Roy, 2023 McGill BIOL200 - Fall 2023 The nuclear pore com...

McGill BIOL200 - Fall 2023 Nucleo-Cytoplasmic Transport © R. Roy, 2023 McGill BIOL200 - Fall 2023 Nuclear pore complexes resemble baskets, and are the sites of nucleocytoplasmic transport Cytoplasmic face Nucleoplasmic face © R. Roy, 2023 McGill BIOL200 - Fall 2023 The nuclear pore complex is a highly ordered structure • It’s huge: 125 megadaltons or 30x bigger than a ribosome. • Composed of ~50 (yeast) to ~100 (vertebrates) different proteins. • Molecules up to ~40-60 kDa can freely diffuse through the NPC. Bigger molecules and multimolecular complexes (RNPs) must be transported. © R. Roy, 2023 McGill BIOL200 - Fall 2023 Transport through the NPC depends on reversible hydrophobic interactions FG nucleoporins are hydrophobic and possess disordered domains © R. Roy, 2023 McGill BIOL200 - Fall 2023 Protein import through the NPC • All nuclear proteins are synthesized in the cytoplasm and imported through NPCs. • These proteins contain a nuclear localization signal (NLS). © R. Roy, 2023 McGill BIOL200 - Fall 2023 Addition of an NLS to a cytoplasmic protein targets it to the nucleus pyruvate kinase NLS-pyruvate kinase © R. Roy, 2023 McGill BIOL200 - Fall 2023 Proteins required for nuclear import • Ran: a monomeric G protein that exists in two conformations; one bound to GTP and another when bound to GDP. • Nuclear Transport Receptors: (Importins) these proteins bind to NLS domains present on cargo proteins to facilitate transport through the pore by associating with FG repeats on the nucleoporins. © R. Roy, 2023 McGill BIOL200 - Fall 2023 Mechanism for nuclear import of NLS-containing cargo proteins High concentration Import complex Low concentration Import complex Low concentration RanGTP/Importin complex High concentration RanGTP/Importin complex © R. Roy, 2023 McGill BIOL200 - Fall 2023 Exportin 1 uses a similar Ran-mediated mechanism © R. Roy, 2023 McGill BIOL200 - Fall 2023 Some RNAs are exported through association with Ran • Exportin t functions to export tRNAs. It binds fully processed tRNAs and Ran-GTP and passes through NPCs. The complex dissociates in the cytosol when it interacts with Ran-GAP. • Export of ribosomal subunits requires Ran. • Some specific mRNAs that associate with particular hnRNP proteins (HIV Rev for example) can be exported through association with Ran. • Most mRNAs are exported in a Ran-independent process using an mRNA exporter. © R. Roy, 2023 McGill BIOL200 - Fall 2023 A simple model of mRNP export Transport of the mRNP through the NPC is Ran-independent • Consists of two subunits (NXF1/NXT1). -Together they bind RNA cooperatively with specific mRNP proteins including the SR proteins. -They form a domain that interacts with FG repeats in nucleoporins. © R. Roy, 2023 McGill BIOL200 - Fall 2023 mRNP packaging in Balbiani Rings Insect polytene chromosomes (Balbiani Rings) provide a system where transcription and mRNP export can be microscopically imaged. Chironomous tentans © R. Roy, 2023 McGill BIOL200 - Fall 2023 Model of mRNP export ribosomes Since ribosomes associate with the mRNA while it is still being exported, the 5’ end must be exported first. © R. Roy, 2023 McGill BIOL200 - Fall 2023 Model of mRNP export • Only fully spliced mature mRNAs get exported; mechanisms of this restriction are not fully understood. • This phenomenon may be associated with the activity of a protein Cytoplasmic bound to a nucleoporin remodelling that actively blocks premRNAs from leaving the nucleus RNA Helicase © R. Roy, 2023