Introduction
Inducing and activating plant immune response is one of the most
effective ways to combat plant virus diseases (Hammerschmidt et al.,
2001). Plant virus that has a variety of infection modes, a wide range
of host and a long infection cycle causes a common systemic infection
(Scholthof, 2005, M. et al., 1982). Control of plant virus disease
largely depends on the application of the virus passivation agents.
However, these agents are not able to provide complete control of the
plant viral diseases owing to their ability to partially reduce the
virus infection and inhibit the replication and transfer of the virus in
the plant(Yang et al., 2016, Yamakawa and H., 1998). Inducers with the
ability to activate plant resistance are becoming an increasingly
important way to control plant virus disease. Previous studies showed
that they can significantly induce the expression of resistance-related
genes and enhance the immune response of plants to biotic
stresses(Yamakawa and H., 1998). However, the application of the
existing inducers in the field is limited because their lasting time is
shorter than the existing period of virus infection. In addition, most
of the inducers consist of biological polysaccharides with a structure
that is easily degraded at the complicated environment in the field,
rustling in the reduced efficacy on the control of virus disease (Xiang
et al., 2013, Lin et al., 2006, Xiu et al., 2006, Tang et al., 2012).
Therefore, there is an urgent need to develop a novel inducer agent with
the ability to last the release time of loaded drugs for the field
application.
Calcium ions (Ca2+) as an essential nutrient element,
can promote plant growth, improve photosynthesis, and increase the
synthesis and accumulation of organic matter (Tang et al., 2012, Kwon et
al., 2009). Ca2+ as a second messenger is also
involved in the signal transduction of plant to stress responses
(Batistic and Kudla, 2012, Defalco et al., 2010, Dodd et al.). Previous
studies showed that Ca2+ signals are perceived by four
types of calcium signaling sensor proteins, including calmodulin
(CaM ), calmodulin-like protein (CML ), calcium-dependent
protein kinase (CDPK), and calcium-like protein Calcineurin B-like (CBL)
(Braam and Davis, 1990, Mccormack et al., 2005, Boonburapong and
Buaboocha, 2007). However, CML, CDPK, CBL are found only in plants and
some protists. CMLs , as one of the plant-specific
Ca2+ receptors, participate in plant biological
stress, abiotic stress as well as many developmental
processes(Scrase-Field and Knight, 2003, Bender and Snedden, 2013,
Nakahara et al., 2012). It has been documented that plant defense
response, plant growth and development as well as cytokine regulation,
induce changes in intracellular calcium ion concentration, which results
in the induced expression of CMLs gene (Roberts and Harmon, 1992,
Mcainsh and Pittman, 2009). Accumulated evidence showed that the
up-regulation of CML gene expression is able to significantly
improve the plant resistance to biotic stress (Ma et al., 2008, Bo et
al., 2017). For example, CML41 was found to reduce P.
syringae infection by regulating plasmodesmal closure through mediating
Ca2+ signaling in response to bacterial pathogens(Bo
et al., 2017). Leba et al. showed that CML9 is involved in
plant resistance to pathogenic bacteria through a flagellin-dependent
pathway in plants(Leba et al., 2012). Tobacco calmodulin-like protein,
rgs-CaM, binds to the RNA silencing (RNAi) suppressor of the tobacco
etch virus to inhibit RNAi and increase tobacco resistance to the virus
(Nakahara et al., 2012). CML19 , also known as arabidopsis centrin
2, was found to be regulated by abiotic stress (UV resistance) and play
diverse roles in DNA damage repair (Liang et al., 2006). Therefore, CML
can be regarded as an important target site to stimulate plant defense
response.
Recent studies showed that sustained-release hydrogel as a drug carrier
has attracted more interest in delivering therapeutic agents to control
human diseases(Jeong et al., 1997, Asamura et al., 2010). Advantages of
hydrogel include a prolonged drug release and action time, increased
stability of loaded drugs, and enhanced control efficiency of
disease(Wasikiewicz et al., 2005, Builders et al., 2008, Ali et al.,
2004). In addition, many studies showed that sustained-release hydrogel
exhibits the ability to carry multiple drugs and provide various control
of multiple diseases. For example, porphyrin photosensitizer
sinoporphyrin sodium (DVDMS) and Polylactic acid-copolymerized glycolic
acid (PLGA) are loaded into the sodium alginate-chitosan hydrogel that
provides the dual action of antibacterial and skin regeneration (Mai et
al., 2020). Therefore, the development of dual functions of hydrogel
that could deliver a range of drugs and provide broad-spectrum
resistance to multiple diseases is required.
Our previous studies found that coating the surface of hydrogel with
amino oligosaccharides can prolong the release time of drugs loaded in
the hydrogel (ALA-gel) (Xiang et al., 2019). However, amino
oligosaccharides are highly soluble in water, resulting in the slow
dissolve of the ALA-gel surface and the reduced drug release time from
the ALA-gel. To overcome the drawbacks of ALA-gel and improve the
sustained release of drugs loaded in the hydrogel, a multiple-functional
sodium alginate-lentinan hydrogel (SL-gel) with a dense chitosan shell
(CSL-gel) was developed aiming at improving the control efficacy against
plant virus diseases. As expected, CSL-gel exhibited stable and
sustained LNT release and broad-spectrum anti-virus activities.
According to the RNA-seq, the sustainable and controlled release of
calcium ions activated the expression of CML19 that enhances the
resistance of tobacco against TMV with a lasting time up to 30 days
after CSL-gel treatment.