1.A.120. The Coronaviral Double Membrane Pore Complex (CoV-DMPC) Family
Coronaviral RNA is replicated in double membrane vesicles (DMVs) by the replicatioin organelle (RO), but the RNA must exit the DMV and enter the cytoplasm in order to be translated into protein. Only the +RNA strand and the mRNAs need to be exported. There appears to be a molecular pore complex that spans both membranes of the DMV Wolff et al. 2020). The pore has an internal diameter of about 260 nm (based on cryo EM) and therefore can probably accomodate both ssRNA and dsRNA. It is crown-shaped with the crown projecting into the cytoplasm where it probably interacts with nsp4 and the viral nucleocapsid (N) protein. The virus encodes three integral membrane proteins, nsp3, nsp4 and nsp6. nsp3 is large with 2309 aas and a size of about 222 kDas. Although it has several (up to 6) regions that are hydrophobic enough to span the membrane, Wolff et al. 2020 claim that it has 2 TMSs; still to span two membranes, it may need 4 TMSs. nsp3 is known to interact with nsp4, and this interaction drives membrane pairing and determines DNV biogenesis and morphology. nsp4 is 56 kDa in size and has 4 putative TMSs; nsp6 is 33 kDa in size with 6 putative TMSs (Wolff et al. 2020). nsp3 has a ubiquitin (Ubl1) domain that is believed to modulate RNA exit to the cytoplasm and may interact with the nucleocapsid (N) protein to stabilze the RNA in the cytoplasm, both for translation and for packaging into the viral particles (the virions). There are likely to be additional proteins, possible both viral and host proteins, that are associated with the pore complex, but only these two proteins will be included in TC entry, TC# 1.A.20.1.1. Coronavirus non-structural proteins, Nso3/6, derive from the polyproteins of various coronaviruses, and usually have 200 - 300 aas, often with about 8 TMSs. They have been proteolytically processed from polyproteins of 4000 to 8000 aas. It is not always clear whether coronaviral proteins referred to in the protein databases are the processed or unprocessed products of the polyproteins. Hyperglycemia (high glucose concentrations) potentiates SARS-CoV-2 infection and plays a central role in severe COVID-19 and diabetes comorbidity (Yusuf et al. 2022).
The reaction believed to be catalyzed by the double membrane pore is:
RNA (lumen of the DMV) → RNA (cytoplasm)
References:
The coronaviral double membrane-spanning pore complex in the replication organelle (RO) of the viral double membrane vesicle (DMV). The pore consists of a hexameric complex. The primary constituent of each of the six subunits of the pore is the non-structural protein-3, nsp3, of 2309 aas, which may have 5 TMSs, based on a hydrophathy plot of the protein. These are clustered together about two-thrids of the way towards the C-terminus. It is the core constituent of the pore and has both its N-terminal domain of 160 kDa and a smaller C-terminal domain, which contains a ubiquitin domain (Ubi1) of 12.6 kDa, that together form the prongs of the cytoplasmic crown. nsp3 interacts with nsp4, and this complex is believed to drive membrane pairing and participate in DMV biogenesis (Wolff et al. 2020). It may associate with other viral and host proteins.
nsp3-nsp4 of murine hepatitis virus
Polyprotein 1a of 4018 aas and ~ 10 TMSs.
PP 1a of Canine coronavirus
ORF1a polyprotein, partial of 4345 aas and ~ 7 TMSs. A cell-based system combined with flow cytometry has been used to evaluate antibody responses against SARS-CoV-2 transmembrane proteins in patients with COVID-19 (Martin et al. 2022).
ORF1a polyprotein of Severe acute respiratory syndrome coronavirus 2, SARS-CoV-2
Non structural protein 6 (Nsp6) of 290 aas and 7 - 9 TMSs. If 7 TMSs, they occur in a 3 + 1 + 3 TMS arrangement. If 8 TMSs, they occur in a 3 + 5 TMS arrangement. If 9 TMSs, they are all closely adjacent to each other. Nsp6 is part of the large polyprotein 1a of this virus.
Nsp6 of severe acute respiratory syndrome coronavirus 2
Polyprotein 1a, partial, of 220 aas and 6 or 7 TMSs
PP1a of Porcine deltacoronavirus
Coronavirus Nsp3 (HD2) of 293 aas and 7 TMSs
Nsp3 of Infectious bronchitis virus