Interplay Between the Host Cell Membrane and dengue virus.

Dengue virus (DENV) is an enveloped, positive sense single-stranded RNA virus belonging to the Flavivirus genus. It is the causative agent of dengue fever, dengue haemorrhagic and dengue shock syndrome, with 50 to 100 million people infected annually. The number of cases of dengue fever and dengue haemorrhagic fever are also on the rise.

DENV infection was found to induce the cytoplasmic membrane rearrangements, which is vital for viral replication in host cells.1 In addition, DENV infection has been shown to perturb cellular levels of HMG-CoA-reductase (HMGCR), the rate limiting enzyme in cholesterol biosynthesis.2 However, it is unknown how modulation of cholesterol affects the viral replication since both inhibition of HMGCR and addition of exogenous cholesterol were found to reduce virus titer.2,3 Since cholesterol is a vital component of the cell membrane, the perturbation of HMGCR could be associated with these membrane rearrangements.

Introduction Objectives

Results

Acknowledgements

Conclusion

C6/36 cell line infected with

DENV-2

This study aims to: 1. To identify the specific virus protein inducing replication vesicle formation in the host cell.2. To reveal the key molecules – protein and lipid species – that induce and maintain

replication vesicle formation in the host cell.3. To uncover the viral capacity to interfere with and manipulate basic cellular metabolism,

based on large-scale protein and lipid analysis by mass spectrometry.

Material and Method

UT-1 cells were treated with Compactin

Cellular Lipids, Proteins analysis by Mass Spectrometry

and cholesterol concentration assayed

using Amplex Red Cholesterol Assay kit

(Invitrogen)Vs

Control

Fixe

d fo

r TE

M a

t 12,

24

, 36,

48

and

72 h

r

Future Works

Positive Control Analysis of Cholesterol levels by MS DENV-2 Infection Induced Cellular Changes

References

control c6/36 cell line 24-hour-DENV-2Compactin treated UT-1 cells 12-hour – DENV-2

48-hour – DENV-2 72-hour – DENV-2

Cholesterol

CHO

Control

Drug

0

50

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Cho

lest

erol

(ug/

ml)

Cholesterol Ester

CHO

Control

Drug

0

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Cho

lest

erol

Est

ers

(ug/

ml)

Department Of Biotechnology Indian Academy Centre for Research & P.G. Studies BangaloreEmail Id: rashmirekha978@gmail.com

Rashmirekha Mohapatra1, Sarshad Ibnu Sayed, Selvam Arjunan.

1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0

Time, min

0.0

2.0e4

4.0e4

6.0e4

8.0e4

1.0e5

1.2e5

1.4e5

Inte

nsity

, cps

Cholesterol

Cholesterol Esters

Control

Drug Treated

• DENV-2 infection increased the level of cholesterol in a time-dependent manner.

• DENV-2 infection leads to membrane arrangements, autophagy interaction and lipid droplet

1. S. Welsch et al., Cell Host Microbe 5, 365 (2009).2. C. Rothwell et al., Virology 389, 8 (2009).3. C. J. Lee et al., J Virol 82, 6470 (2008).4. L. M. Pfeffer et al., Proc Natl Acad Sci U S A 82, 2417 (1985).5. J. Y. Luu et al., Hum Pathol 20, 617 (1989).

1. To verify the key molecule (proteins/lipids) in the induced membrane formation after DENV-2 infection in c6/36 cell lines.

2.To elucidate change in cholesterol localization induced by DENV-2.

I would like to express my gratitude to the UGC for funding and National Institute of virology for providing technical assistance on laboratory and NIMHANS for electron Microscopy. I would also like to thank the members of department of biotechnology of Indaian Academy college for their support.