In contrast to the defence mechanism of RNA interference (RNAi) in plants and invertebrates, its role in the innate response to virus infection of mammals is a matter of debate. Since RNAi has a well-established role in controlling infection of the alphavirus Sindbis virus (SINV) in insects, we have used this virus to investigate the role of RNAi in SINV infection of human cells

A Chakrabarti

A Mortazavi

A Saumet

Andras Donaszi-Ivanov

BC Ho

BR Cullen

BR tenOever

CL Campbell

CM Cirimotich

CW Burke

E Gottwein

EG Strauss

EM Morazzani

ER Mardis

EY Choy

F Ma

F Weber

G Gatto

G Szittya

I Mohorianu

IP Greene

Irina Mohorianu

J Fang

JI Henke

JK Ahluwalia

JR Abend

JY Leung

K Prufer

KJ Ishii

KL McKnight

KM Myles

KW Witwer

L Du

M Hariharan

MB Stocks

MC Saleh

ML Metzker

MP Gantier

N Vodovar

P Parameswaran

Penny P. Powell

PV Maillard

RL Pilcher

RL Skalsky

RP Kincaid

RW Williams

S Griffiths-Jones

S Koyama

S Moxon

SW Ding

T Kawai

Tamas Dalmay

V Stollar

W Hou

WB Klimstra

X Lei

Y Kim

Y Li

YQ Wu

Zach N. Adelman

ZN Adelman

English

PubMed

In contrast to the defence mechanism of RNA interference (RNAi) in plants and invertebrates, its role in the innate response to virus infection of mammals is a matter of debate. Since RNAi has a well-established role in controlling infection of the alphavirus Sindbis virus (SINV) in insects, we have used this virus to investigate the role of RNAi in SINV infection of human cells.SINV AR339 and TR339-GFP were adapted to grow in HEK293 cells. Deep sequencing of small RNAs (sRNAs) early in SINV infection (4 and 6 hpi) showed low abundance (0.8%) of viral sRNAs (vsRNAs), with no size, sequence or location specific patterns characteristic of Dicer products nor did they possess any discernible pattern to ascribe to a specific RNAi biogenesis pathway. This was supported by multiple variants for each sequence, and lack of hot spots along the viral genome sequence. The abundance of the best defined vsRNAs was below the limit of Northern blot detection. The adaptation of the virus to HEK293 cells showed little sequence changes compared to the reference; however, a SNP in E1 gene with a preference from G to C was found. Deep sequencing results showed little variation of expression of cellular microRNAs (miRNAs) at 4 and 6 hpi compared to uninfected cells. Twelve miRNAs exhibiting some minor differential expression by sequencing, showed no difference in expression by Northern blot analysis.We show that, unlike SINV infection of invertebrates, generation of Dicer-dependent svRNAs and change in expression of cellular miRNAs were not detected as part of the Human response to SINV

Penny P Powell

Directory of Open Access Journals

PLoS ONE

Small RNA analysis in Sindbis virus infected human HEK293 cells.

In contrast to the defence mechanism of RNA interference (RNAi) in plants and invertebrates, its role in the innate response to virus infection of mammals is a matter of debate. Since RNAi has a well-established role in controlling infection of the alphavirus Sindbis virus (SINV) in insects, we have used this virus to investigate the role of RNAi in SINV infection of human cells.Deep sequencing results showed little variation of expression of cellular microRNAs (miRNAs) at 4 and 6 hpi compared to uninfected cells. Twelve miRNAs exhibiting some minor differential expression by sequencing, showed no difference in expression by Northern blot analysis.We show that, unlike SINV infection of invertebrates, generation of Dicer-dependent svRNAs and change in expression of cellular miRNAs were not detected as part of the Human response to SINV

Donaszi-Ivanov Andras

Mohorianu Irina

Dalmay Tamas

Powell Penny P.

Public Library of Science (PLOS)

Small RNA Analysis in Sindbis Virus Infected Human HEK293 Cells

<div>IntroductionIn contrast to the defence mechanism of RNA interference (RNAi) in plants and invertebrates, its role in the innate response to virus infection of mammals is a matter of debate. Since RNAi has a well-established role in controlling infection of the alphavirus Sindbis virus (SINV) in insects, we have used this virus to investigate the role of RNAi in SINV infection of human cells.ResultsSINV AR339 and TR339-GFP were adapted to grow in HEK293 cells. Deep sequencing of small RNAs (sRNAs) early in SINV infection (4 and 6 hpi) showed low abundance (0.8%) of viral sRNAs (vsRNAs), with no size, sequence or location specific patterns characteristic of Dicer products nor did they possess any discernible pattern to ascribe to a specific RNAi biogenesis pathway. This was supported by multiple variants for each sequence, and lack of hot spots along the viral genome sequence. The abundance of the best defined vsRNAs was below the limit of Northern blot detection. The adaptation of the virus to HEK293 cells showed little sequence changes compared to the reference; however, a SNP in E1 gene with a preference from G to C was found.Deep sequencing results showed little variation of expression of cellular microRNAs (miRNAs) at 4 and 6 hpi compared to uninfected cells. Twelve miRNAs exhibiting some minor differential expression by sequencing, showed no difference in expression by Northern blot analysis.ConclusionsWe show that, unlike SINV infection of invertebrates, generation of Dicer-dependent svRNAs and change in expression of cellular miRNAs were not detected as part of the Human response to SINV.</div

Andras Donaszi-Ivanov (503936)

Irina Mohorianu (503937)

Tamas Dalmay (251047)

Penny P. Powell (253815)

The Francis Crick Institute

Crossref

Donaszi-Ivanov, Andras

Mohorianu, Irina

Dalmay, Tamas

Powell, Penny P

University of East Anglia digital repository

Small RNA analysis in Sindbis virus infected human HEK293 cells

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http://dx.doi.org/10.1371/journal.pone.0084070

Small RNA analysis in Sindbis virus infected human HEK293 cells

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