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1.A.57 The Human SARS Coronavirus Viroporin (SARS-VP) Family

The severe acute respiratory syndrome-associated coronavirus group 26 (SARS CoV) genome includes Orf3a which codes for a protein, Protein 3a (Protein IX; Protein U276; Acessory Protein 3a) of 274 aas that has 3 putative N-terminal TMSs (residues 1-130) and a C-terminal hydrophilic domain (residues 131-274). It is expressed and is found at the surface of SARS-CoV-infected or 3a-cDNA transfected cells (Lu et al, 2006). It forms a homotetrameric (composed of two homodimers) K+-sensitive channel, with intersubunit disulfide bridges. It is inhibited by Ba2+. It appears to be important for release of the virus from the cell but not for replication of the viral DNA (Lu et al, 2006). Protein 3a may be glycosylated (as in the virion) or nonglycosylated.

Accessory protein 3a interacts with several other SARS CoV proteins. These include protein E, protein-7A, the spike and M proteins and the human small glutamine-rich tetratricopeptide repeat (TPR)-containing protein A [(O43765; 313 aas)].  Viroporins have been reviewed with respect to their structures, functions, and their roles in the life cycle of SARS-CoV-2 (Breitinger et al. 2022).

The SARS CoV envelope small membrane protein E (E protein, sM protein (P59637)) of 76 aas with 1 TMS is the prototype of small homologues (70-90 aas) found in several coronaviruses specific for rats, cats, dogs, mice, people, cows, etc. These include transmissible gastroenteretis virus, mouse hepatitis virus, rat sialodacryoadenitis coronavirus, porcine hemagglutinating encephalomyelitis virus.

The Protein 7a precursor (P59635; protein X4; protein U122; accessory protein 7a) with 122 aas, has 1 TMS and is found in the endoplasmic reticulum/golgi apparatus connecting region as well as in the virion.

Fourteen ORFs are encoded in the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) genome (Lu et al. 2006). This family of proteins has been referred to as the DUF3092 family. Emodin inhibits current through SARS-associated coronavirus 3a protein (Schwarz et al. 2011). The ion channel activity of the 3a protein is linked to its pro-apoptotic function (Chan et al. 2009).

Protein 3a is a 274 amino acid polytopic channel protein with three TMSs encoded by SARS-CoV. Synthetic peptides corresponding to each of its three individual TMSs have been reconstituted into artificial lipid bilayers (Chien et al. 2013). Only TMSs 2 and 3 induced channel activity. Reconstitution of the peptides as TMS1 + TMS3 or TMS2 + TMS3 in a 1: 1 mixture induced membrane activity. In a 1: 1: 1 mixture, channel-like behavior was almost fully restored. Expression of full length 3a and reconstitution into artificial lipid bilayers revealed a weakly cation-selective (PK ≈ 2 PCl ) rectifying channel. In the presence of non-physiological concentrations of Ca2+-ions, channel activity developed.

The sequences of three polytopic viral channel proteins: (i) p7 of HCV and 2B of Polio virus (two TMSs) and (ii) 3a of SARS-CoV (three TMSs were aligned, and the region of overlap was mapped onto structural models of host channels and toxins, focusing on the pore-lining TMSs (Schindler and Fischer 2012). The analysis reveals that p7 and 2B TMSs align with the pore-facing TMS of MscL, and 3a-TMSs align with those of ligand-gated ion channels (Schindler and Fischer 2012). 

Plant synaptotagmins (SYTs) are resident proteins of the endoplasmic reticulum (ER) (Benitez-Fuente and Botella 2023). They are characterized by an N-terminal transmembrane region and C2 domains at the C-terminus, which tether the ER to the plasma membrane (PM). In addition to their tethering role, SYTs contain a lipid-harboring SMP domain, essential for shuttling lipids between the ER and the PM. There is evidence on Arabidopsis SYT1, the best-characterized family member (see TC#s 9.A.57.1.5 and 1.7), which link it to biotic and abiotic responses as well as to ER morphology. Benitez-Fuente and Botella 2023 reviewed knowledge of SYT members, focusing on their roles in stress, and they discuss how these roles can be related to their tethering and lipid transport functions. These authors also contextualize this information about SYTs with their homologs, the yeast tricalbins and the mammalian extended synaptotagmins.

The reaction catalyzed by SARS-VP is:

Small molecules (in) small molecules (out)

This family belongs to the: Viroporin-2.

References associated with 1.A.57 family:

Ambrożek-Latecka, M., P. Kozlowski, G. Hoser, M. Bandyszewska, K. Hanusek, D. Nowis, J. Gołąb, M. Grzanka, A. Piekiełko-Witkowska, L. Schulz, F. Hornung, S. Deinhardt-Emmer, E. Kozlowska, and T. Skirecki. (2024). SARS-CoV-2 and its ORF3a, E and M viroporins activate inflammasome in human macrophages and induce of IL-1α in pulmonary epithelial and endothelial cells. Cell Death Discov 10: 191. 38664396
Barrantes, F.J. (2021). Structural biology of coronavirus ion channels. Acta Crystallogr D Struct Biol 77: 391-402. 33825700
Benitez-Fuente, F. and M.A. Botella. (2023). Biological roles of plant synaptotagmins. Eur J. Cell Biol. 102: 151335. [Epub: Ahead of Print] 37390668
Bianchi, M., A. Borsetti, M. Ciccozzi, and S. Pascarella. (2020). SARS-Cov-2 ORF3a: Mutability and function. Int J Biol Macromol. [Epub: Ahead of Print] 33359807
Breitinger, U., N.S. Farag, H. Sticht, and H.G. Breitinger. (2022). Viroporins: Structure, function, and their role in the life cycle of SARS-CoV-2. Int J Biochem. Cell Biol. 145: 106185. [Epub: Ahead of Print] 35219876
Chan, C.M., H. Tsoi, W.M. Chan, S. Zhai, C.O. Wong, X. Yao, W.Y. Chan, S.K. Tsui, and H.Y. Chan. (2009). The ion channel activity of the SARS-coronavirus 3a protein is linked to its pro-apoptotic function. Int J Biochem. Cell Biol. 41: 2232-2239. 19398035
Chen, I.Y., M. Moriyama, M.F. Chang, and T. Ichinohe. (2019). Severe Acute Respiratory Syndrome Coronavirus Viroporin 3a Activates the NLRP3 Inflammasome. Front Microbiol 10: 50. 30761102
Chien, T.H., Y.L. Chiang, C.P. Chen, P. Henklein, K. Hänel, I.S. Hwang, D. Willbold, and W.B. Fischer. (2013). Assembling an ion channel: ORF 3a from SARS-CoV. Biopolymers 99: 628-635. 23483519
Fam, M.S., C.A. Sedky, N.O. Turky, H.G. Breitinger, and U. Breitinger. (2023). Channel activity of SARS-CoV-2 viroporin ORF3a inhibited by adamantanes and phenolic plant metabolites. Sci Rep 13: 5328. 37005439
Gruner, H.N., Y. Zhang, K. Shariati, N. Yiv, Z. Hu, Y. Wang, J.F. Hejtmancik, M.T. McManus, K. Tharp, and G. Ku. (2023). SARS-CoV-2 interacts with the Clic-like chloride channel-1 () and triggers an unfolded protein response. PeerJ 11: e15077. 37033725
Henke, W., M. Kalamvoki, and E.B. Stephens. (2023). The Role of the Tyrosine-Based Sorting Signals of the ORF3a Protein of SARS-CoV-2 on Intracellular Trafficking, Autophagy, and Apoptosis. bioRxiv. 37547007
Kern, D.M., B. Sorum, S.S. Mali, C.M. Hoel, S. Sridharan, J.P. Remis, D.B. Toso, A. Kotecha, D.M. Bautista, and S.G. Brohawn. (2021). Cryo-EM structure of SARS-CoV-2 ORF3a in lipid nanodiscs. Nat Struct Mol Biol 28: 573-582. 34158638
Lu, W., B.J. Zheng, K. Xu, W. Schwarz, L. Du, C.K. Wong, J. Chen, S. Duan, V. Deubel, and B. Sun. (2006). Severe acute respiratory syndrome-associated coronavirus 3a protein forms an ion channel and modulates virus release. Proc. Natl. Acad. Sci. USA 103: 12540-12545. 16894145
Nie, Y., L. Mou, Q. Long, D. Deng, R. Hu, J. Cheng, and J. Wu. (2023). SARS-CoV-2 ORF3a positively regulates NF-κB activity by enhancing IKKβ-NEMO interaction. Virus Res 328: 199086. 36894068
Schindler, C. and W.B. Fischer. (2012). Sequence alignment of viral channel proteins with cellular ion channels. J Comput Biol 19: 1060-1072. 22891808
Schwarz, S., K. Wang, W. Yu, B. Sun, and W. Schwarz. (2011). Emodin inhibits current through SARS-associated coronavirus 3a protein. Antiviral Res 90: 64-69. 21356245
Tee, W.V., Z.W. Tan, K. Lee, E. Guarnera, and I.N. Berezovsky. (2021). Exploring the Allosteric Territory of Protein Function. J Phys Chem B 125: 3763-3780. 33844527
Zhang, B., Y. Jin, L. Zhang, H. Wang, and X. Wang. (2022). Pentamidine Ninety Years on: the Development and Applications of Pentamidine and its Analogs. Curr. Med. Chem. [Epub: Ahead of Print] 35289252