Cotton leaf curl Burewala Virus complex: Virus-Host interactions.

Department of Bioinformatics and BiotechnologyComputer Science

Govt. College University, Faisalabad

GOVERNEMNET COLLEGE UNIVERSITY, FAISALABAD

Abstract

Existing fair calculation methods for resetting the target in interrupted cricket Formatted: Indent: First line: 0.5", No bullets or

matches are claimed by most of the researchers to be fair but these are biased towards either team
Formatted: Default Paragraph Font, Font: (Default) Times
batting first or team batting second. People think that existing method does not consider all the New Roman, 12 pt, Pattern: Clear (White)
important factors which effect the target score and are only consider some of the factor, which are Formatted: Default Paragraph Font, Font: (Default) Times
New Roman, 12 pt, Pattern: Clear (White)
the main cause of creating bias. In this research, a new algorithm is proposed which will consider
Formatted: Line spacing: Multiple 1.15 li
these factor like innings momentum, historical scoring patterns and players rating. Data mining is
Formatted: Default Paragraph Font, Font: (Default) Times
used to get historical scoring patterns. New Roman, 12 pt, Pattern: Clear (White)
Formatted: Default Paragraph Font, Font: (Default) Times
Searching, ranking and prediction of domain experts or contents has gained the attention of the New Roman, 12 pt, Pattern: Clear (White)
researchers with the advent of online social networks and online databases. Several works are put Formatted: Default Paragraph Font, Font: (Default) Times
New Roman, 12 pt, Pattern: Clear (White)
forward by different experts to address these issues.
Formatted: Default Paragraph Font, Font: (Default) Times
New Roman, 12 pt, Pattern: Clear (White)
 Future prediction of citation count and temporal expert finding are such proposal among
many other results. But these proposals did not consider sports domain in appropriate manner.
While considering the sports domain, some issues related to sports forecasting are discussed in but
proper prediction mechanism is not presented by the authors. Hence WEKA machine learning
library is considered to predict match in terms of different aspects. Moreover, none among above
proposals considered cricket domain. Taking cricket game into an account. There are further
proposals for ranking of batsman based on performance metric for ranking of cricket teams by
employing h-index and Page Ranking. However, the aforementioned proposals will not only
predict the match status but also the performance of player as well.Cotton leaf curl disease
(CLCuD) is the major plant viral constraint to cotton production on the Asian subcontinent.
CLCuD is primarily caused by begomovirus, Cotton leaf curl Burewala virus (CLCuBuV), and
associated satellite Cotton leaf curl Multan betasatellite (CLCuMB). Begomovirus is the largest
genus among geminiviruses having more than 200 species and transmitted by whitefly (Bemisia
tabaci) vector. Cotton leaf curl Burewala virus (CLCuBuV) is a whitefly-transmitted monopartite
begomovirus and has recombinant genome composed of sequences derived from CLCuMuV
and Cotton leaf curl Kokhran virus (CLCuKoV) that were associated with cotton leaf curl disease
(CLCuD) during 1990s. The genome of monopartite begomovirus is organized into six ORFs viz.
AC1 (Rep), AC2 (TrAp), AC3 (REn), AC4, AV1 (CP) and AV2 which are transcribed bidirectionally.
Most monopartite begomoviruses are associated with satellite molecule called betasatellite which
is often required for the development of typical symptoms in naturally-infected host plant. It
requires a helper begomovirus for its replication, systemic infection and whitefly vector-mediated
transmission. In the present study I will clone all the genes of Cotton leaf curl Burewala virus and
associated satellite in plant expression binary vector under CaMV-35 promoter. Then viral genes
will be expressed in GFP transgenic Nicotiana benthamiana 16c line along with GFP RNAi
construct to check the suppression ability of proteins encoded by Cotton leaf curl Burewala
complex. Then small RNA profiling of GFP gene and viral genes will be done to analyze the
suppression efficiency of each gene. Further viral proteins will be expressed in bacterial system
and purified using 6X-Histidin tag and interactions of purified viral proteins with different host
factors involved in host defense mechanism will be done.
 GOVERNEMNET COLLEGE UNIVERSITY, FAISALABAD

Cotton leaf curl Burewala Virus complex: Virus-Host interactions.

Abstract

Existing fair calculation methods for resetting the target in interrupted cricket matches are
claimed by most of the researchers to be fair but these are biased towards either team batting first
or team batting second. People think that existing method does not consider all the important
factors which effect the target score and are only consider some of the factor, which are the main
cause of creating bias. In this research, a new algorithm is proposed which will consider these
factor like innings momentum, historical scoring patterns and players rating. Data mining is used
to get historical scoring patterns.

Searching, ranking and prediction of domain experts or contents has gained the attention of the
researchers with the advent of online social networks and online databases. Several works are put
forward by different experts to address these issues.
Future prediction of citation count and temporal expert finding are such proposal among
many other results. But these proposals did not consider sports domain in appropriate manner.
While considering the sports domain, some issues related to sports forecasting are discussed in but
proper prediction mechanism is not presented by the authors. Hence WEKA machine learning
library is considered to predict match in terms of different aspects. Moreover, none among above
proposals considered cricket domain. Taking cricket game into an account. There are further
proposals for ranking of batsman based on performance metric for ranking of cricket teams by
employing h-index and Page Ranking. However, the aforementioned proposals will not only
predict the match status but also the performance of player as well.Cotton leaf curl disease
(CLCuD) is the major plant viral constraint to cotton production on the Asian subcontinent.
CLCuD is primarily caused by begomovirus, Cotton leaf curl Burewala virus (CLCuBuV), and
associated satellite Cotton leaf curl Multan betasatellite (CLCuMB). Begomovirus is the largest
genus among geminiviruses having more than 200 species and transmitted by whitefly (Bemisia
tabaci) vector. Cotton leaf curl Burewala virus (CLCuBuV) is a whitefly-transmitted monopartite
begomovirus and has recombinant genome composed of sequences derived from CLCuMuV
and Cotton leaf curl Kokhran virus (CLCuKoV) that were associated with cotton leaf curl disease
(CLCuD) during 1990s. The genome of monopartite begomovirus is organized into six ORFs viz.
AC1 (Rep), AC2 (TrAp), AC3 (REn), AC4, AV1 (CP) and AV2 which are transcribed bidirectionally.
Most monopartite begomoviruses are associated with satellite molecule called betasatellite which
is often required for the development of typical symptoms in naturally-infected host plant. It
requires a helper begomovirus for its replication, systemic infection and whitefly vector-mediated
transmission. In the present study I will clone all the genes of Cotton leaf curl Burewala virus and
associated satellite in plant expression binary vector under CaMV-35 promoter. Then viral genes
will be expressed in GFP transgenic Nicotiana benthamiana 16c line along with GFP RNAi
construct to check the suppression ability of proteins encoded by Cotton leaf curl Burewala
complex. Then small RNA profiling of GFP gene and viral genes will be done to analyze the
suppression efficiency of each gene. Further viral proteins will be expressed in bacterial system
and purified using 6X-Histidin tag and interactions of purified viral proteins with different host
factors involved in host defense mechanism will be done.

Cotton leaf curl disease (CLCuD) is the major plant viral constraint to cotton production
on the Asian subcontinent. CLCuD is primarily caused by begomovirus, Cotton leaf curl Burewala
virus (CLCuBuV), and Cotton leaf curl Multan betasatellite (CLCuMB). Begomovirus is the
largest genus among geminiviruses having more than 200 species and transmitted by whitefly
(Bemisia tabaci) vector. Cotton leaf curl Burewala virus (CLCuBuV) is a whitefly-transmitted
monopartite begomovirus and has recombinant genome composed of sequences derived from
CLCuMuV and Cotton leaf curl Kokhran virus (CLCuKoV) that were associated with cotton leaf
curl disease (CLCuD) during 1990s. The genome of monopartite begomovirus is organized into
six ORFsviz.AC1 (Rep), AC2 (TrAp), AC3 (REn), AC4, AV1 (CP) and AV2 which are transcribed
bidirectionally. Most monopartite begomoviruses are associated with satellite molecule called
betasatellite which is often required for the development of wild type symptom in naturally-
infected host plant. It requires a helper begomovirus for its replication, systemic infection and
whitefly vector-mediated transmission. In the present study I will clone all the genes of Cotton leaf
curl Burewala virus and associated satellite in plant expression binary vector under CaMV-35
promoter. Then viral genes will be expressed in GFP transgenic Nicotiana benthamiana 16c line
along with GFP RNAi construct to check the suppression ability of proteins encoded by Cotton
leaf curl Burewala complex. Then small RNA profiling of GFP gene and viral genes will be done
to analyze the suppression efficiency of each gene. Further viral proteins will be expressed in
bacterial system and purified using 6X-Histidin tag and interactions of purified viral proteins with
different host factors involved in host defense mechanism will be done.

Key Words: begomovirusData Mining; betasatelliteResetting Targets in Cricket Matches; post-

transcriptional gene silencingforecasting; RNA silencing suppressorFair Calculation, Predictive
Algorithm
 Introduction

Nevertheless, very little efforts are made towards providing efficient mechanisms for
ranking teams, or to find experts from the teams. Recently, some social networking analysts
among scholastic communities have attracted towards this dimension due to its over grown
popularity. Working towards this direction, Brace-well & Ruggiero utilized parametric control
chart for performance monitoring of a batsman in different matches [3]. Amin & Sharma
employed ordered weighted averaging technique coupled with regression structure in order
to measure the performance, and to rank T20 batsmen.

Cricket game prediction is one, who currently own relatively weak profiles, but can be
predicted as the future experts of the respective domains. Rising Star Prediction (RSP) is made
based on the current contributions of RSs coupled by considering their ascending performance
and joint collaborations with the domain experts. Finding such RSs within the organizational
domains is the great need of current era, so that the organizations can put efforts to maximize the
expertise of RSs in order to get the optimal performance in future.

Hence, RSP is an emerging research dimension that inspires us to forecast RSs from
sports domain. Considering the sports domain, among all sports, cricket is the second most
popular game that was originated from England, and now has its roots round the globe.
Therefore, we take cricket game as our case study for RSP. The players in cricket game can be
categorized into different classes based on their performance evolution. This is all because cotton
leaf curl disease and its diverse strains.

In South Asia, Cotton leaf curl disease (CLCuD) is caused by a complex of

phylogenetically-related begomovirus species and a specific betasatellite that is transmitted by the
whitefly Bemisia tabaci . This disease was first reported in 1967 on upland cotton (Gossypium
hirsutum L.) at Khokhran near Multan on few individual plants (Saeed et al., 2012; Hussain & Ali,
1975). It became a serious problem after 1991-92 and continued till the development of Cotton
Leaf Curl resistant variety in 1996. But this problem arose again during 2001 in Burewala breaking
the resistance of all the available cotton cultivars/lines (Mansoor et al., 2003). Now cotton leaf curl
virus is one of the most destructive disease of cotton in the Punjab area of Pakistan. This virus is
8% different from the previous virus on molecular basis (Mansoor et al., 2003), but symptoms are
almost the same to the previous CLCuV.

The physical characteristics of the cotton leaf curl disease are upward curling of leaves, the
veins of the affected leaves become thickened which are most pronounced on the underside. Two
types of veins thickenings are observed, small vein thickening and main vein thickening. Initial
symptoms consisted for transient vein clearing on young leaves (Mansoor et al., 1993; Nateshan
et al., 1996). In extreme but not infrequent cases, plants became stunted and give fewer yields.
Many workers have reported that due to cotton leaf curl virus yield has been decreased greatly
(Harrison et al., 1997; Brown, 2001; Ahmad et al., 2002).

The viruses associated with the CLCuD complex on the Asian subcontinent, are single
component begomoviruses (genus Begomovirus family Geminiviridae ). The satellite (DNA β)
and satellite-like (DNA 1) components have yet to be classified, although the DNA 1 components
are closely related to, and thought to have originated from, components of a second group of single-
stranded DNA viruses, the nanoviruses (family Nanoviridae ). Physical properties of
begomoviruses associated with CLCuD, like all geminiviruses, have geminate (twinned) particles,
approximately 18–20 nm in diameter and 30 nm long, consisting of two incomplete T = 1
icosahedra joined together in a structure with 22 pentameric capsomers and 110 identical protein
subunits. It is probable, although not conclusively proven, that the DNA 1 and DNA β components,
being half the size of the viral component, are encapsidated in monomeric, rather than geminate
particles (Briddon 2003).

The ability of a plant virus to effectively infect a particular host depends on the balance
between host defenses and virus multiplication. Among host defense responses, RNA silencing
has emerged as an important antiviral mechanism in plants. RNA silencing refers to a homology-
dependent gene-silencing mechanism guided by small RNAs (18 to 25 nucleotides nt long),
including short interfering RNAs (siRNAs) and microRNAs (miRNAs). Upon infection, plants
can process viral RNA into siRNAs and use them to direct specific antiviral silencing activity.
Several lines of evidence, especially from studies into siRNAs and viral pathogenesis, have led to
the current view of cytoplasmic RNA silencing, generally referred to as post-transcriptional gene
silencing (PTGS), as a ubiquitous defense against RNA viruses and transcripts produced by DNA
viruses (Ana et al., 2012).
 Many viruses express products that actively block the function of the antiviral RNAi
pathway, termed viral suppressors of RNAi (VSRs) or RNA Silencing Suppressors (RSSs). They
are found in RNA and DNA viruses, with both plant and animal hosts. Suppression of RNAi
pathway b VSR may often be a key stage of viral infection and some viruses even encode multiple
VSRs (e.g potyviruses; P1 and Hc-Pro). (Gemma et al., 2013). The ability of the begomovirus
Cotton leaf curl Multan virus (CLCuMV) and its associated betasatellite, Cotton leaf curl Multan
ß-satellite (CLCuMB) which, together, cause CLCuD, and the nonessential alphasatellite (Cotton
leaf curl Multan alphasatellite [CLCuMA]) for their ability to suppress gene silencing in Nicotiana
benthamiana has been observed and analyzed (Amin et al., 2011).

The long-term approach to cope with this problem and to save the cotton crop from the
ravages of CLCuV is the development of Cotton Leaf Curl resistant varieties (Akhtar et al., 2002),
as previously practiced in Sudan and Egypt (Kirkpatrick, 1931; Khan et al., 2001).

The new strain of virus known as Cotton leaf curl burewala virus (CLCuBuV) started
damaging the cotton crop in many areas of Pakistan during 2001. All the cotton leaf curl resistant
commercial varieties showed susceptibility to CLCuBuV. The symptoms and alternate hosts of
CLCuBuV are similar to those of previous cotton leaf curl virus. After emergence of this new
strain of virus the yield loss started increasing. The hot spots of this disease increased from 2.4%
to 73.4% during the year 2001-2007. Similarly the area affected under this disease also increased
from 4.45% to 70.26% during this period. At present no single variety is resistant to CLCuBuV.
This is alarming that the hot spots as well as area affected by this disease are increasing every
year and the situation is becoming verse. So the aims and objectives of this study is to find the
suppressors of the CLCuBuV for the development of resistance varities.

Aims and Objectives

 Cloning of all the genes of Cotton leaf curl Burewala virus and associated satellite.

 Identification of Suppressors by inoculating all cloned genes in Nicotiana

Benthamiana16c line along with GFP RNAi

 siRNA profiling GFP gene after suppressor identification

 Recombinant expression of viral protein in Bacterial sytem E.Coli

  In-vitro binding assays of proteins encoded by CLCBuV complex with different Species
of RNA molecules

MATERIALS AND METHOD

To work on the project following methodology will be used

1. Primer Designing:
Primers will be designed on the basis of available sequence (GenBank) of cotton leaf curl
burewala virus and its beta satellite with the help of software available.

2. Cloning of all genes of CLCuBuV:

All the genes of Cotton leaf curl Burewala virus and associated satellite will be PCR amplified.
The amplicons will be purified and restricted by appropriat restriction enzymes. The restricted
products will be ligated into pJIT163 under 35 promoter and cloned into E.Coli Top10.
.
3. Sequencing and sequence analysis of Samples

Confirmed clones will be sent for DNA sequencing to the company which provide DNA
sequencing facility commercially and sequences will be analysed by using different bioinformatics
tools.

4. Development of RNAi construct

RNAi constructs will be developed by lifting the cloned genes along with 35S promoter from
pJIT163 to the binary expression vector pGreen 0029 .That construct will express the ds RNA to
initiate RNA silencing mechanism .

5. Expression of RNAi construct in Model plants:

RNAi construct will be transformed into Agrobacterium tumefaciens strain GV3101 and
inoculated into model plants Nicotiana benthamiana to check the RNAi GFP activity.

6. Evaluation of RNAi based resistance :

Analysis of suppression activity of individual gene will be done in RNAi transgenic plants along
with non-transgenic plants by visual detection of GFP under UV light.

7. Expression of Protein :

All the genes will be expressed in bacterial system using pQE30 vector under T5 promoter. The
viral proteins will purified using His-Tag and binding affinity of purified protein with different
species of RNA molecule (ssRNA, ds RNA, short RNA and long RNA) will be analyzed.

Review of Literature

Plant viruses are the most troublesome and a great challenge in the world of phytopathogens, as
they cause devastating diseases of economically important crops (Hull and Davies, 1992). Plant
viruses are classified into 49 families and 73 genera having more than 600 species infecting a
very broad range of important crops. The first ever virus to be discovered was the plant virus
Tobacco mosaic virus (TMV)., as they cause devastating diseases of economically important
crops (Hull and Davies, 1992). Plant viruses are classified into 49 families and 73 genera having
more than 600 species infecting a very broad range of important crops. The first ever virus to be
discovered was the plant virus Tobacco mosaic virus (TMV). The discovery of TMV is often
credited to Martinus Beijerinck who determined, in 1898, that plant sap obtained from tobacco
leaves with "mosaic disease" remained infectious when passed through a porcelain filter (Lecoq,
2001).

Viruses of the family Geminiviridae have circular single-stranded (ss) DNA genomes that are
encapsidated in characteristic twinned icosahedral (geminate) capsids (Hanely et al., 2013). They
are classified into seven genera i.e Begomovirus, Curtovirus, Mastrevirus, Topocovirus,
Becurtovirus, Eragrovirus, Truncurtovirus (Adams et al., 2013 and Brown et al., 2012). The most
abundant geminiviruses are those that are transmitted by the whitefly Bemisia tabaci and are
classified in the genus Begomovirus.

A majority of the begomoviruses originating from the New World (NW) have genomes consisting
of two components, known as DNA A and DNA B, both of which are required for virus infectivity.
Recently, a single monopartite begomovirus (with a genome consisting of only a homolog of DNA
A components of bipartite viruses) has been identified in South America (Melgarejo et al., 2013). In
the Old World (OW), however, although a small number of bipartite viruses have been identified,
the majority of begomoviruses have monopartite genomes.

Satellites are defined as subviral agents composed of nucleic acids and that depend on co-infection
with a HV for their multiplication (Briddon et al., 2008). Betasatellites are single stranded circular
DNA molecules exclusively associated with monopartite begomoviruses (family Geminiviridae).
Betasatellites depend on the helper begomoviruses for their replication and encapsidation and
enhance virus accumulation in the host plants. Betasatellites are about half of the size of their
helper begomoviruses (~1350 nt). They contain an adenine rich (A-rich) region, a sequence that is
highly conserved between all betasatellites, known as the “satellite conserved region (SCR), which
is about 150-200 nt in length. Betasatellites encode a single protein on the complementary-sense
strand known as βC1 (Briddon et al., 2008). It is responsible for symptoms induction (Briddon and
Stanley, 2006).

Alphasatellites are associated with some monopartite begomovirus-betasatellite complexes. They

are called “satellite-like” molecules because they are self replicating but depend on helper
begomovirus for encapsidation, and whitefly transmission; by definition, satellites depend upon
their HV for replication. The precise function of begomovirus-associated alphasatellites is still
unclear since experimentally they are not essential for infection or disease development (Briddon
et al., 2004). It is observesd that two alphasatellites encode Rep proteins with suppressor of PTGS
activity. It is thus possible that alphasatellites contribute to overcoming host defenses (Nawaz-ul-
Rehman et al., 2010).

The genomes of monopartite, and DNA A components of bipartite, begomoviruses encode in the
virion-sense the coat protein (CP) which is required for encapsidation, transmission by insects, and
movement in plants and the V2 protein, homologous to the AV2 protein for bipartite
begomoviruses, is involved in virus movement and may act as an RSS. The complementary-sense
strand encodes (i) the replication-associated protein (Rep); a rolling-circle replication initiator
protein; (ii) the transcriptional-activator protein (TrAP) which is involved in the up-regulation of
late viral gene expression, modulates host gene expression and suppresses gene silencing; (iii) the
replication enhancer protein (REn) which is involved in creating a cellular environment that is
conducive to virus replication and (iv) the C4 protein which may be a pathogenicity determinant
and an RSS (Fondong. 2013 and Sharma et al., 2008).

RNA silencing, a natural defense mechanism evolved to protect eukaryotic genomes against
invasive nucleic acids such as transgenes, transposable elements and viruses, has emerged as a
powerful tool with a wide range of applications in functional genomics and genetic engineering
(Baulcombe, 2004; Vaucheret, 2006; Voinnet, 2008). RNA silencing was first discovered in plants
as „co-suppression‟, where the expression of both an introduced transgene and the homologous
endogenous chalcone synthase gene were co-suppressed (Napoli et al., 1990). RNA level is
reduced in RNA silencing, by inhibition of transcription (transcriptional gene silencing [TGS]), by
activation of sequence-specific RNA degradation (post-transcriptional gene silencing [PTGS]) and
DNA degradation (Sidahmed and Wilkie, 2010). Sequence-specific RNA degradation is referred
to as RNA interference (RNAi) in animals, as PTGS in plants and as quelling in fungi (Baulcombe,
2004; Baulcombe, 2002; Baulcombe, 2006; Dykxhoorn et al., 2003).

PTGS is a sequence-specific RNA degradation process that occurs in the cytoplasm and leads to
the accumulation of 21 to 24 nt-long siRNAs corresponding to the silenced gene (Fagard and
Vaucheret, 2000; Vaucheret et al., 2001:). PTGS can be induced by viruses, sense transgenes,
antisense transgenes and hairpin-RNA-expressing transgenes with inverted repeats. The dsRNA
trigger is processed into 21 to 24 nt long siRNAs (Hamilton and Baulcombe, 1999) by a
ribonuclease III type enzyme known as Dicer, which was initially identified in Drosophila
(Bernstein et al., 2001; Tang et al., 2003; Zamore et al., 2000). Dicers are large (~200 kDa),
multidomain proteins that contain a putative RNA helicase domain, a PAZ
(Piwi/Argonaute/Zwille) domain, two tandem ribonuclease III (RNase III) domains and one or two
dsRNA-binding domains. A. thaliana encodes four Dicer-like proteins and each has specialized
functions (Schauer et al., 2002; Xie et al., 2004). The DICER-LIKE (DCL) endonucleases, DCL2,
DCL3, and DCL4 process dsRNA into 22-, 24-, and 21-nucleotides (nt) long small interfering
RNAs (siRNAs), respectively (Fusaro et al., 2006).

The silencing mechanism by which the DNA sequence of gene is modified in such a way that
transcription is inhibited, is known as transcriptional gene silencing (TGS) (Chandler and
Vaucheret, 2001; Matzke and Birchler, 2005). Transcriptionally silenced genes have inactive
promoters (Baulcombe, 2004; Baulcombe, 2002). TGS occurs in the nucleus and is generally
associated with methylation of promoter sequences (Fagard and Vaucheret, 2000). DNA
methylation is induced by RNA signals which come from various sources, including invasive viral,
transgene and transposon sequences and in some cases from ssRNA precursors produced by host
plant RDR.

RNA silencing is used by plants as an antiviral defense system. Viruses have evolved various
silencing suppression strategies to counter this defense mechanism (Baulcombe, 2004; Carrington
et al., 2001; Voinnet et al., 1999). The presence of virus-encoded silencing suppressors was
indicated by the observation that the severity of disease caused by other viruses increased in the
presence of potyviruses (Mlotshwa et al., 2002). Subsequently it was shown that the Helper
Component-Protease (HC-Pro) of potyviruses is the factor medaiting this synergism (Pruss et al.,
1997), which is due to interference in the silencingmediated antiviral defense (Anandalakshmi et
al., 1998). Viral Suppressors of RNA silencing interfere at different steps of RNA silencing
(Omarov and Scholtho 2012). Some bind dsRNA and sequester siRNAs away from the RNAi
pathway, Binds to Ago, preventing the RNA-induced silencing complex (RISC) from cleaving
target RNA, degradation of Ago, Others inhibit cell-to-cell signaling, some even encode multiple
supperssors. (Burgyan 2008; Voinnet 2005). Some the VSRs and their functions are as followed:
The post-transcriptional gene silencing (PTGS) suppression activities of the transcriptional
activator protein (TrAP), C4, V2 and βC1 proteins encoded by Cotton leaf curl Kokhran virus
(CLCuKoV)/CLCuMuB have been assessed in Nicotiana benthamian and observed the variable
degree of silencing suppression, with V2 protein exhibiting the strongest suppression activity and
only the C4 protein preventing the spread of systemic silencing. The CLCuKoV-encoded TrAP,
C4, V2 and CLCuMuB-encoded βC1 proteins. TrAP shown to bind various small and long nucleic
acids including single-stranded (ss) and double-stranded (ds) RNA and DNA molecules. C4, V2,
and βC1 bound ssDNA and dsDNA with varying affinities (Saeed et al., 2015).

VIRUS VSR FUNCTION REFERENCE

turnip mosaic virus
Lakatos et al., 2005
(TurMV) P1and HcPro siRNA and signal
Kasschau et al., 2003
potato virus Y (PVY)

potato virus X (PVX) P25 Argonaute and signal Bayne et al., 2005 Chiu et al., 2010

tomato-spotted wilt
NSs siRNA Bucher et al., 2003
virus (TSWV)

sugarcane yellow leaf

P0 Argonaute Mangwende et al., 2003
virus (SYLV)

rice yellow mottle

P1 siRNA Lacombe et al., 2010
virus (RYMV)

tobacco mosaic virus

P30 signal Kubota et al., 2003
(ToMV)
cucumber mosaic virus Brigneti et al., 1998
2b Argonaute
(CMV) Canto et al., 2001
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