Pandemic Origin Biology Unit 3: Molecular Genetics

Lecture 3.7: SARS-CoV-2 and COVID-19

John D. Nagy

BIO 181: General Biology for Majors, Scottsdale Community College

2020 Revision

John Nagy Lec 3.7: SARS-CoV-2 1/26 Pandemic Origin Biology Outline

1 Current Status of the Pandemic

2 Origin of the Virus

3 Basic Biology of Coronaviruses Anatomy Genome Life Cycle

John Nagy Lec 3.7: SARS-CoV-2 2/26 Pandemic Origin Biology Difference between SARS-CoV-2 and COVID-19

The UN’s World Health Organization is responsible for officially designating the the disease and its causative agent [7].

John Nagy Lec 3.7: SARS-CoV-2 3/26 Pandemic Origin Biology Early dynamics of the outbreak

Data through March 2. Blue: Number of cases. Red: Number of deaths. Source: Rabi et al. 2020 [8].

John Nagy Lec 3.7: SARS-CoV-2 4/26 Pandemic Origin Biology Current status of the epidemic: November 6, 2020

From the Johns Hopkins Coronavirus Resource Center: coronavirus.jhu.edu/data/new-cases

John Nagy Lec 3.7: SARS-CoV-2 5/26 Pandemic Origin Biology SARS-CoV-2 did not come from nowhere

Many coronaviruses infect hu- mans: Original sources are bats and rodents. First 4 listed cause mild disease (common colds). SARS-CoV (2003-04) and MERS-CoV (2012) caused severe epidemics. Most (all?) have secondary hosts from which they infected humans. Sources: Corman et al. 2018 [3]; Cui et al. 2019 [4].

John Nagy Lec 3.7: SARS-CoV-2 6/26 Pandemic Origin Biology Origins of 2 other serious coronavirus diseases

Definition A zoonotic disease can be transmitted from other species to humans. SARS-CoV:

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Bats Civets Humans MERS-CoV:

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Bats Dromedary camels Humans John Nagy Lec 3.7: SARS-CoV-2 7/26 Pandemic Origin Biology Clues to the source of SARS-CoV-2

Left: Entire genome compar- ison. Right: Comparison of a single gene, RNA-dependent RNA polymerase (RdRp).

Discovery: Zhang et al. 2020 [12] Pangolins carry CoV that is genetically very similar to human SARS-CoV-2.

John Nagy Lec 3.7: SARS-CoV-2 8/26 Pandemic Origin Biology More clues to the source of SARS-CoV-2

Amino acid sequence comparison (from Zhang et al. 2020 [12]): Gray: A critical region of the protein that allows virus to enter cells. Yellow: Deletion in most bat CoVs. Orange: Amino acids (residues) that bind to ACE2 on host cells—allows virus to enter cells.

Conclusion Pangolin CoV is identical to human SARS-CoV-2 at critical binding sites (orange); most similar bat CoV is not.

John Nagy Lec 3.7: SARS-CoV-2 9/26 Pandemic Origin Biology Evidence so far

Pangolin-CoV is about 91% identical to SARS-CoV-2. Bat RaTG13, SARS-CoV-2 & pangolin-CoV extremely similar. Pangolin-CoV & SARS-CoV-2-have similar binding motifs.

SARS-CoV-2:

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Bats Pangolins Humans John Nagy Lec 3.7: SARS-CoV-2 10/26 Pandemic Origin Biology Probable origin of SARS-CoV-2

Further observations (evidence not shown): SARS-CoV-2 doesn’t resemble laboratory SARS (Andersen et al. 2020 [1]). Most of the original cases had visited the Huanan market in Wuhan, China [8, 11, 13]. Genetic analysis Conclusions suggests the virus entered the human SARS-CoV-2 probably originated at a population in market in Wuhan, China that sold November or early pangolins for meat. December, 2019 [1].

John Nagy Lec 3.7: SARS-CoV-2 11/26 Pandemic Origin Biology Anatomy Genome Life Cycle Basic anatomy of SARS-CoV

Viral genes are coded on one, single-stranded RNA molecule in each virion (Stadler et al. 2003 [10]). Key proteins: Spike (S): cell entry. Envelope (E): embedded in membrane derived from host cell. Nucleocapsid (N): Protects viral genome.

John Nagy Lec 3.7: SARS-CoV-2 12/26 Pandemic Origin Biology Anatomy Genome Life Cycle Genome of SARS-CoV

Original SARS-CoV genome is well-understood (Stadler et al. 2003 [10]). Replicase/transcriptase complex (ORF1 1a/1b) includes genes for replicating the virus. The 30 end houses genes for the spike, envelope, nucleocapsid and other structural proteins.

1ORF = “open reading frame” John Nagy Lec 3.7: SARS-CoV-2 13/26 Pandemic Origin Biology Anatomy Genome Life Cycle Comparison of human severe coronaviruses

SARS-CoV-2 is genetically similar to SARS-CoV and MERS-CoV (Li et al. 2020 [6]).

John Nagy Lec 3.7: SARS-CoV-2 14/26 Pandemic Origin Biology Anatomy Genome Life Cycle Key aspects of SARS-CoV-2 genome

SARS-CoV-2 translation (Cascella et al. 2020 [2]).

John Nagy Lec 3.7: SARS-CoV-2 15/26 Pandemic Origin Biology Anatomy Genome Life Cycle General coronvirus life cycle

S, E, and M proteins are built in the rough ER. Embedded in membrane there. N proteins are made in the cytoplasm. Assembly thought to occur between RER and Golgi. Assembly completed in Golgi. Virus is shed via exocytosis. What about SARS-CoV-2?

John Nagy Lec 3.7: SARS-CoV-2 16/26 Pandemic Origin Biology Anatomy Genome Life Cycle Life cycles of SARS-CoV and MERS-CoV

Source: Song et al. 2019 [9].

John Nagy Lec 3.7: SARS-CoV-2 17/26 Pandemic Origin Biology Anatomy Genome Life Cycle What about SARS-CoV-2?

Research question: how does SARS-CoV-2 enter cells?

Hypotheses The spike protein is almost certainly involved. But does it bindn to ACE2 like SARS-CoV does? DPP4 like MERS-CoV does? Something novel?

John Nagy Lec 3.7: SARS-CoV-2 18/26 Pandemic Origin Biology Anatomy Genome Life Cycle Comparison of spike primary structure

Observation The primary structure of the receptor-binding domain of S is like that of coronaviruses that use ACE2 (Hoffmann et al. 2020 [5]).

John Nagy Lec 3.7: SARS-CoV-2 19/26 Pandemic Origin Biology Anatomy Genome Life Cycle Receptor binding domain of SARS-CoV

Source: Cascella et al. 2020 [2].

John Nagy Lec 3.7: SARS-CoV-2 20/26 Pandemic Origin Biology Anatomy Genome Life Cycle Testing viral entry against ACE2 and other proteins

Bars show how many viruses enter cells. VSV-G is the positive control. Dark and light blue are ACE2-expressing cells. Green is DPP4. Lower case, “h” means “human.” (Hoffmann et al. 2020 [5]).

John Nagy Lec 3.7: SARS-CoV-2 21/26 Pandemic Origin Biology Anatomy Genome Life Cycle Another important part of the story

Bars and VSV-G same as previous. TMPRSS2 cleaves ACE2. E-64d is a drug that inhibits cleavage by enzymes other than TMPRSS2 (Hoffmann et al. 2020 [5]).

Do these results support, contradict, or say nothing about the hypothesis that TMPRSS2 is required to SARS-CoV-2 to infect cells?

John Nagy Lec 3.7: SARS-CoV-2 22/26 Pandemic Origin Biology Anatomy Genome Life Cycle Our current understanding

Source: Rabi et al. 2020 [8].

Current hypothesis of host cell entry SARS-CoV-2 infection cycle: S protein binds to host cell ACE2; host cell TMPRSS2 cleaves ACE2, which activates the S protein; after activation, virus can enter cell and begin infection.

John Nagy Lec 3.7: SARS-CoV-2 23/26 Pandemic Origin Biology Anatomy Genome Life Cycle ReferencesI

Kristian G. Andersen, Andrew Rambaut, W. Ian Lipkin, Edward C. Holmes, and Robert F. Garry. The proximal origin of SARS-CoV-2. Nat. Med., Online, 2020.

Marco Cascella, Michael Rajnik, Arturo Cuomo, Scott C. Dulebohn, and Raffaela Di Napoli. Features, evaluation and treatment coronoavirus (COVID-19). In StatPearls. StatPearls, Treasure Island, FL, Updated April 6 2020.

Victor M. Corman, Doreen Muth, Daniela Niemeyer, and Christian Drosten. Hosts and sources of endemic human coronaviruses. Adv. Virus Res., 100, 163–188 2018.

Jie Cui, Fang Li, and Zheng-Li Shi. Origin and evolution of pathogenic coronaviruses. Nat. Rev. Microbiol., 17:181–192, 2019.

Markus Hoffmann, Hannah Kleine-Weber, Simon Schroeder, Nadine Kr¨uger,Tanja Herrler, Sandra Erichsen, Tobias S. Schiergens, Georg Herrler, Nai-Huei Wu, Andreas Nitsche, marcel A. M¨uler, Christian Drosten, and Stefan P¨ohlmann. SARS-CoV-2 cenn entry depends on ACE2 and TMPRSS2 and is blocked by a clinically proven protease inhibitor. Cell, 181:In press, 2020.

Xiaowei Li, Manman Geng, Yizhao Peng, Liesu Meng, and Shemin Lu. Molecular immune pathogenesis and diagnosis of COVID-19. J. Pharm. Analysis, In press, 2020.

John Nagy Lec 3.7: SARS-CoV-2 24/26 Pandemic Origin Biology Anatomy Genome Life Cycle ReferencesII

World Health Organization. Naming the coronavirus disease (COVID-19) and the virus that causes it (www.who.int/emergencies/diseases/novel-coronavirus-2019/technical-guidance/naming- the-coronavirus-disease-(covid-2019)-and-the-virus-that-causes-it), April 2020. Firas A. Rabi, Mazhar S. Al Zoubi, Ghena A. Kasasbeh, Dunia M. Salameh, and Amjad D. Al-Nasser. SARS-CoV-2 and coronavirus disease 2019: What we know so far. Pathogens, 9:231, 2020.

Zhiqi Song, Yangeng Xu, Linlin Bao, Ling Zhang, Pin Yu, Yajin Qu, Hua Zhu, Wenjie Zhao, Yunlin Han, and Chuan Qin. From SARS to MERS, thrusting coronaviruses into the spotlight. Viruses, 11:59, 2019.

Konrad Stadler, Vega Masignani, Markus Eickmann, Staphan Becker, Sergio Abrignani, Hans-Dieter Klenk, and Rino Rappuoli. SARS—beginning to understand a new virus. Nat. Rev. Microbiol., 1:209–218, 2003.

Fan Wu, Su Zhao, Bin Yu, Yan-Mei Chen, and et al. A new coronavirus associated with human respiratory disease in China. Nature, 579:265–269, 2020.

Tao Zhang, Qunfu Wu, and Zhigang Zhang. Probable pangolin origin of SARS-CoV-2 associated with the COVID-19 outbreak. Curr. Biol., 30:1346–1351, 2020.

John Nagy Lec 3.7: SARS-CoV-2 25/26 Pandemic Origin Biology Anatomy Genome Life Cycle ReferencesIII

Peng Zhou, Xing-Lou Yang, Xian-Guang Wang, Ben Hu, and et al. A pneumonia outbreak associated with a new coronavirus of probably bat origin. Nature, 579:270–273, 2020.

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