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MICaB Faculty

Leslie A. Schiff
Leslie A. Schiff

Professor

Department of Microbiology

Tufts University, 1985, Ph.D.

612-624-9933 - office
612-626-0623 - fax

E-mail:schif002@umn.edu


Research Interests:

Virus-host interactions, viral entry, translational control in virus-infected cells, viral immune avoidance mechanisms

Reovirus Life Cycle

Diagram of the reovirus life cycle

Overview:

Mammalian orthoreoviruses are prototypical members of the family Reoviridae, which includes the pathogenic rotaviruses, coltiviruses and orbiviruses. Orthoreoviruses (reoviruses) are poorly pathogenic in humans. This characteristic, together with their preferential replication in transformed cells, has led to their recent development as oncolytic agents for the treatment of human tumors. However, despite the ongoing trials of reovirus therapy in humans, relatively little is known about the molecular determinants (viral and cellular) that regulate infection. My laboratory uses mammalian reoviruses to understand molecular interactions that influence the outcome of viral infection. Our studies have focused on functions of sigma 3, one of the most abundant proteins in the reovirus virion (see cryo-EM diagram below) and reovirus-infected cell. Sigma 3 functions in two distinct, yet fundamentally important steps in the viral life cycle: penetration of the viral particle into the cell cytoplasm and the regulation of protein synthesis/interferon-resistance in the infected cell.

Reovirus TIL Surface Shaded

Surface-shaded views of reovirus TIL (left) showing a split particle composed of a TIL viron and an ISVP and (right) a split particle composed of a T1L ISVP and a core particle. Pictures are based on three-dimensional reconstruction of data obtained by cryoelectron microscopy. Image provided by Dr. Max Nibert, Harvard Medical School (my favorite collaborator!)

Space Filling Sigma 3

Space filling diagram of sigma 3 showing conserved and non-conserved residues. Residues in red are not conserved among different serotypes and isolates. Conserved residues are in gray.Views (A) and (B) are rotated by 180 degrees with respect to each other about the long axis. Residues thought, based on mutational analysis, to be involved in interactions with mu 1 are shown in blue. Lys 293, thought to be involved in the binding of dsRNA, is shown in yellow. The protease-hypersensitive loop is at the bottom right of (A) and at the top right of (B). From; Olland, A.M., Jane-Valbuena, J., Schiff, L.A., Nibert, M.L., and Harrison, S.C. (2001) Structure of the reovirus outer capsid protein sigma 3 at 1.8 angstroms resolution. EMBO J. 20(5):1-11.

Detailed Information:

Research Area 1

Research Area 2

Relevant Publications

External Research Support

Search Entrez Browser for Schiff L

Professional Experience

Honors

Professional Affiliations

Current Teaching

A Few Favorite Links

Personal Interests

 

Research Area 1:

Unlike other enteric viruses that typically require extracellular proteolysis to initiate infection, reoviruses appear to exploit both intracellular and extracellular enzymes to degrade sigma 3 and prepare the particle for viral entry. Our recent work has focused on how proteolysis influences the capacity of reovirus to infect cells and cause disease. Joe Golden discovered that the requirement for sigma 3 removal represents a significant host-range determinant in reovirus infection. He found that many cells that otherwise support efficient reovirus replication, restrict infection at this step in the lifecycle. Joe’s thesis work went on to demonstrate that reovirus strains differ in their susceptibility to uncoating by specific proteases and that this can impact cellular host range. This work ultimately contributed to the important finding that reoviruses replicate preferentially in tumor cells because they are more efficiently uncoated in cultured tumor cells and the in vivo tumor microenvironment. Joe’s work also showed that distinct proteases mediate uncoating in different cell types. Studies in cells that express the cysteine protease cathepsin S or the inflammatory serine protease elastase support a model in which the pH-sensitivity of reovirus infection reflects the acid-dependence of particular proteases involved in uncoating, rather than an requisite inherently acid-sensitive step in viral entry.

RA1

Our present goals are to identify cellular proteases and viral determinants of proteolysis that regulate reovirus cell entry and to understand how these factors impact infection and pathogenesis in the host. We are using genetic and biochemical approaches to identify sequences within capsid protein sigma 3 that regulate cell entry. We are selecting and characterizing viral mutants with expanded protease sensitivity and we will use infectious particles recoated with recombinant sigma 3 and a newly developed reverse genetic system to identify determinant of protease susceptibility crucial for cell entry. Within the next year, we plan to move our work into animals, using well-developed mouse infection models and knockout mice to investigate the role that specific intracellular and extracellular proteases play in regulating reovirus tropism, spread and disease. Our studies of the molecular determinants of reovirus cell entry will provide fundamental information concerning the biology of non-enveloped viruses and will facilitate the safe and effective use of reoviruses as oncolytic agents to treat human tumors.

Research Area 2:

The second line of research in my laboratory addresses the fundamental question of how viruses modulate the cellular translation machinery to favor their own replication. We are investigating both the mechanisms by which reovirus shuts off host cell translation and the mechanisms that enable viruses to replicate in the face of global host shutoff. Based on a genetic polymorphism between two prototypic reovirus strains, it has long been hypothesized that the interferon-induced eIF2 alpha kinase, PKR, is responsible for reovirus-induced host shutoff.

However Jen Smith’s thesis work revealed that, while PKR may be entirely responsible for host shutoff after infection with the prototypic strain Jones, other molecules contribute to this phenotype in cells infected with most other reovirus strains. Her infection studies in knockout cells revealed that RNase L, another interferon-induced antiviral molecule, contributes to reovirus-induced host shutoff.

S35 Profile

Total profile of 35S-methionine-labeled proteins in reovirus-infected (D,87,J) or uninfected (mock or M) L929 cells. The positions of viral proteins are indicated at the right of each figure. Notice that host protein synthesis is dramatically inhibited after infection with reovirus clone 87 or Jones (J). Host translational shutoff is minimal after infection with strain Dearing (D). The difference in the host translation profiles between the Jones infection and the Dearing infection is determined by the gene encoding the viral protein sigma 3.

 

Interestingly, while these molecules inhibit host protein synthesis in infected cells, they do not significantly restrict viral replication. In fact, virus yields can be higher in the presence of these ‘antiviral’ molecules! To gain further insight into the causes and consequences of reovirus-induced host shutoff and to better understand how reovirus counters these cellular defenses, we performed a comparative microarray analysis with our colleague Paul Bohjanen (manuscript in preparation). We found that the cellular responses to infection vary quite dramatically depending upon the infecting strain. Interestingly, infection with virus strains that induce host shutoff alters the expression of genes involved in the cellular integrated stress response. These include PERK, an ER resident PKR-like eIF2 alpha kinase and P58IPK, a cellular inhibitor of PKR and PERK. An important goal in the near term is to understand how reovirus infection activates cellular stress pathways. Another important goal is to understand the mechanisms that enable reovirus mRNAs to be efficiently translated in cells in which PERK and PKR phosphorylate and inactivate the eukaryotic translation initiation factor eIF2 alpha . One hypothesis, emanating from the work of Stephen Schmechel in the lab, is that sigma 3 locally inhibits dsRNA-activated molecules such as PKR and RNAse L, to spare translation in areas of viral replication (see figure below). Over the last thirty years, significant insight into translational regulation has been gained from studies of virus-infected cells. Our studies in reovirus-infected cells are likely to provide new and important information regarding mechanisms of eukaroytic translation initiation under conditions of cellular stress.

Localized perinuclear expression of sigma 3 in reovirus c87-infected cells.

Relevant Publications:

  • Schiff, L (2008). Structure and Functions of the Orthoreovirus sigma 3 Protein. In: Segmented Double-Stranded RNA viruses: Structure and Molecular Biology, Patton, ed. Calister Academic Press.
  • Goodman, A.G., Smith, J.A., Balachandran, S., Perwitawari, O., Proll, S.C., Thomas, M.J., Korth, M.J., Barber, G.N., Schiff, L.A. and Katze, M.G. (2007). The cellular protein P58IPK regulates influenza virus mRNA translation and replication through a PKR-mediated mechanism. J. Virol. 81(5): 2221-2230. (selected for inclusion in JVI Spotlight)
  • Alain, T., Kim, TSY, Lun XQ, Liacin A, Schiff LA, Senger DL, Forsyth PA (2007) Proteolytic Disassembly is a Critical Determinant for Reovirus Oncolysis. Mol Ther. [Epub ahead of print]
  • Smith JA, Schmechel SC, Raghavan A, Abelson M, Reilly C, Katze MG, Kaufmam RJ, Bohjanen PR and Schiff LA (2006) Reovirus induces and benefits from a cellular integrated stress response. J. Virol. Feb;80(4):2019-2033. Abstract JVI Spotlight
  • Schiff, L, Nibert, M.L. and Tyler, K. (2006) Reoviruses. In: Fields Virology. Knipe, Howley, eds. LWW, publishers.
  • Smith JA, Schmechel SC, Williams BR, Silverman RH, Schiff LA (2005) Involvement of the interferon-regulated antiviral proteins PKR and RNase L in reovirus-induced shutoff of cellular translation. J Virol. Feb;79(4):2240-50. Abstract
  • Golden JW, Schiff LA (2005). Neutrophil elastase, an acid-independent serine protease, facilitates reovirus uncoating and infection in U937 promonocyte cells.
    Virol J. May 31;2:48 Abstract
  • Nibert ML, Odegard AL, Agosto MA, Chandran K, Schiff LA (2005) Putative autocleavage of reovirus mu1 protein in concert with outer-capsid disassembly and activation for membrane permeabilization. J Mol Biol. 2005 Jan 21;345(3):461-74. Abstract
  • Golden JW, Bahe JA, Lucas WT, Nibert ML, and Schiff LA (2004) Cathepsin S. Supports Acid-independent Infection by Some Reoviruses. J Biol Chem. Mar 5;279(10):8547-5). Abstract
  • Golden JW, Linke J, Schmechel SC, Thoemke K and Schiff LA (2002) Addition of exogenous protease facilitates reovirus infection in many restrictive cells. J. Virol. 76(15):7430-7443. Abstract
  • Jane-Valbuena J, Breun LA, Schiff LA, and Nibert ML (2002) Sites and determinants of early cleavages in the proteolytic processing pathway of reovirus surface protein sigma 3. J. Viriol 76(10):5184-5197. Abstract
  • Olland AM, Jane-Valbuena J, Schiff LA, Nibert ML, and Harrison SC (2001) Structure of the reovirus outer capsid protein sigma 3 at 1.8 angstroms resolution. EMBO J. 20(5):1-11. Abstract
  • Schiff LA and Nibert ML (2001) Reoviruses and their replication In: Virology. D.M. Knipe, R. Lamb, P. Howley eds. Lippincott Williams & Wilkins, Inc.
  • Gillian AL, Schmechel SC, Livny J, Schiff LA, and Nibert ML (2000) Reovirus nonstructural protein sigmaNS binds in multiple copies to single-strand RNA molecules and exhibits ATP-independent duplex-unwinding activity. J. Virol. 74(13): 5939-5948. Abstract
  • Jane-Valbuena J, Nibert ML, Walker SB, Baker TS, Chen Y, Centonze VE and Schiff LA (1999) Reovirus virion-like particles obtained by recoating ISVPs with baculovirus-expressed sigma3 protein: an approach for analyzing sigma3 functions during virus entry. J. Virol. 73(4): 2963-2973. Abstract
  • Chandran K, Walker SB, Chen Y, Contreras CM, Schiff LA, Baker TS, and Nibert ML (1999) Infectious virion-like particles generated by recoating reovirus cores with baculovirus- expressed mu1 and sigma3 proteins. J. Virol. 73(5): 3941-3950. Abstract
  • Schiff LA(1998) Reovirus outer capsid proteins sigma3 and mu1: Interactions that influence viral entry, assembly and translational control. In: Current Topics in Microbiology and Immunology. Vol. 233/ 1 pp. 167-183. K.L Tyler and M.B.A. Oldstone, eds. Springer Verlag.
  • Schmechel S, Anderson R, Chute M, Meurs E, Skinner P and Schiff L (1997) Preferential translation of reovirus mRNA by a sigma3-dependent mechanism. Virology, 232 (1): 62-73. Abstract
  • Shepard DA, Ehnstrom JG, Skinner PJ and Schiff L (1996) Mutations in the zinc-binding motif of the reovirus capsid protein sigma3 eliminate its ability to associate with capsid protein mu1. J. Virol. 70:2065-2068. Abstract
  • Nibert M, Schiff LA and Fields BN (1996) Reoviruses and their replication. In: Virology. B. Fields, D.M. Knipe, R.M. Chanock, M. Hirsch, J.L. Melnick, T. Monath, B. Roizman, eds. Raven Press, Inc.
  • Kedl R, Schmechel S, and Schiff L (1995) Comparative sequence analysis of the reovirus S4 gene from 13 serotype 1 and serotype 3 field isolates. J. Virol. 69: 552-559. Abstract
  • Shepard, D., Ehnstrom, J., and Schiff, L (1995) Association of reovirus outer capsid proteins sigma3 and mu1 renders sigma3 protease sensitive. J. Virol. 69: 8180-8184. Abstract
  • Nibert ML, Schiff LA, and Fields BN (1991) Mammalian reoviruses contain a myristoylated structural protein. J. Virol. 65:1960-1967. Abstract
  • Dermody TS, Schiff LA, Nibert M., Coombs KM, and Fields BN (1991) The S2 gene nucleotide sequences of prototype strains of the three reovirus serotypes:Characterization of reovirus core protein sigma2. J. Virol. 65:5721-5731. Abstract
  • Schiff LA and Fields BN (1990). Reoviruses and their replication. In: Virology. B. Fields, D.M. Knipe, R.M. Chanock, M. Hirsch, J.L. Melnick, T. Monath, B. Roizman, eds. Raven Press, Inc.
  • Schiff LA, Nibert ML, Co, MS, Brown EG, and Fields BN(1988). Distinct binding sites for zinc and double-stranded RNA in the reovirus outer capsid protein sigma3. Mol. Cell. Biol. 8:273-283. Abstract
  • Schiff LA, Nibert ML, and Fields BN(1988). Characterization of a zinc blotting technique: Evidence that a retroviral gag protein binds zinc. Proc. Natl. Acad. Sci. USA 85:4195-4199. Abstract

 

External Research Support:

  • NIH:Structure and function of reovirus major outer capsid proteins. 1992-1997.
  • ACS:Functional analysis of a viral dsRNA-binding protein.1998-1999.
  • NIH: Reovirus sigma 3 protein: structural and regulatory functions.1999-2010.

 

Present and Past Trainees:

Current Ph.D. Trainees

Rachel Nygaard (on the left)

Rachael and Linse

Wade Schulz

Wade Schulz

 

Past Ph.D. Trainees:

Jennifer Smith, Ph.D.
Currently a postdoctoral fellow with P. Howley, Harvard Medical School

Jennifer Smith

Joseph Golden, Ph.D.
Currently a postdoctoral fellow at USAMARIID

Joseph Golden

Stephen Schmechel, M.D/Ph.D.
Currently an Assistant Professor, Department of Laboratory Medicine and Pathology, University of Minnesota

Stephen Schmechel

 

Former undergraduates who have pursued directed research in the lab:

Ross Kedl, Ph.D. (Asst. Professor, Dept. of Immunology, UCHSC), Brent Huberty (Manager, Pharmacia/Upjohn Sterility Testing Laboratory), Pamela Skinner, Ph.D. (Asst. Professor, University of Minnesota Veterinary School), Robert Anderson, M.D./Ph.D. (ASM undergraduate research fellowship awardee), Michael Chute (Manager, BSL3 Laboratory, Biological Defense Research Directorate, Naval Medical Research Center), Jeffrey Ehnstrom, M.D., Kara Thoemke, Ph.D. (Asst. Professor, College of St. Scholastica), Jay Santos (ASM undergraduate research fellowship awardee), Ashley Fuller, MD (OB/Gyn resident, UT-Southwestern), Patrick Laitala (Advanced Microbial Systems), Tseganesh Selameab, M.D. (resident, Boston Medical Center), Amanda Kostyk, Ph.D. (M.D./Ph.D. University of Colorado), Jessica Linke (National Conference on Undergraduate Research participant; Biology teacher, Delano High School), Eva Chung (Mentored through the Presidents Distinguished Faculty Mentor Program, Ph.D. student in Immunology at Duke), Daniel Westholm (Ph.D. candidate, University of Minnesota, Duluth), Brian Finstead (Beckman-Coulter, Production Scientist), Taj Melson (medical student at University of Michigan), Linse Lahti (Junior Scientist University of Minnesota)

Jessie Bahe
Jessie Bahe (currently a high school science teacher)

Linse Lahti Linse LahtiLinse Lahti (Junior Scientist and Superscuba girl)

 

Professional Experience:

  • Teaching Assistant, Tufts University, 1983
  • Teaching Assistant, Harvard Medical School, 1986
  • Postdoctoral Associate, Harvard Medical School, 1985-1990
  • Assistant Professor of Microbiology, University of Minnesota, 1990-1996
  • Associate Professor of Microbiology,University of Minnesota, 1996-present
  • Director of Undergraduate Studies in Microbiology, Univ. of Minnesota, 1999-present
  • Professor of Microbiology, University of Minnesota, 2005 - present

 

Honors:

  • National Institutes of Health Predoctoral Fellowship, 1979
  • National Institutes of Health Postdoctoral Fellowship, 1987-1989
  • University of Minnesota Presidents Distinguished Mentor Award, 2000
  • University of Minnesota Center for Interdisciplinary Study of Writing Award for Excellence in the Teaching of Writing, 2000
  • Morse-Alumni Award for Outstanding Contributions to Undergraduate Education 2001-2002
  • American Society for Microbiology: Carski Foundation Distinguished Teaching Award, 2004
  • Convocation Speaker, University of Minnesota, 2003
  • Committee on Institutional Cooperation Academic Leadership Fellow, 2007-2008

 

Professional Affiliations:

  • American Society for Microbiology, 1981-present
  • American Society for Virology, 1989-present

 

Current Teaching:

Current Teaching at the Undergraduate Level

  • MicB 4141: Biology, Genetics and Pathogenesis of Viruses (Writing Intensive), 2000-present
  • MicB 4171: Biology, Genetics and Pathogenesis of Viruses (non-writing intensive), 2004-present

 

Recent Teaching at the Graduate Level

  • MIMP 8002: Structure, Function, and Genetics of Bacteria and Viruses, 2000-2005
  • MICaB 8010: Microbial Pathogenesis, 2001-present
  • Grad 8101: Teaching in Higher Education (graduate and post-graduate level), 2002
  • Grad 8200: Active Learning and Course Design in the Sciences (graduate and postgraduate level), 2004

Presenter at teaching enhancement workshops sponsored by the University of Minnesota Center for Teaching and Learning Services and the Center for Writing:

Less Yields More: Using Short Writings in Writing Intensive Courses, Winter 2000 and Fall 2001; Using Technology to Enhance Learning and Teaching Objectives, Spring 2001; Enhancing Large Lecture Courses: A Showcase, Fall 2001; Enhancing Large Lecture Courses: Continuing the Discussion, Fall 2001; Making Use of Short and Informal Writing Assignments, Fall 2002, Fall 2003, and Fall 2004; Teaching Writing One-on-One, Fall 2003; Grading Writing, Fall, 2003; Writing-to-Learn, Fall 2003: Teaching with Writing Online, Fall, 2005; Designing writing assignments that promote learning, Fall 2005; Incorporating writing instruction into “content” courses, Spring 2006; Teaching with Writing Online and Grammar matters, Fall 2006; What’s Grammar Got To Do With It, Spring 2007.

Favorite Links:

 

Personal Interests:

  • Cooking
  • Canine Agility and Obedience

 

Huey Jose and Steve
Huey (yellow australian shepherd), Jose (long-haired german shepherd) and Steve (herpesvirologist and co-inhabitant of lab space in 1425 Mayo)

Noah Reading
Noah Hai Lam Rice (doing some reading in his stroller while visiting the lab

Noah
Noah Hai Lam Rice (5 years old, kidratee)

Noah and Nikki
Noah and friend Nikki, Nature of Life peer mentors, 2022

Lab on Bike

 

Last updated: October 30, 2007