Ectopic phytocystatin expression increases nodule numbers and influences the responses of soybean (Glycine max) to nitrogen deficiency

Abstract

Keywords:Cystatin Cysteine protease Lateral root density Leaf senescence Nitrogen deficiency Nodulation Vacuolar processing enzymes abstract Cysteine proteases and cystatins have many functions that remain poorly characterised, particularly in crop plants. We therefore investigated the responses of these proteins to nitrogen deficiency in wild-type soy-beans and in two independent transgenic soybean lines (OCI-1 and OCI-2) that express the rice cystatin, oryzacystatin-I (OCI). Plants were grown for four weeks under either a high (5 mM) nitrate (HN) regime or in the absence of added nitrate (LN) in the absence or presence of symbiotic rhizobial bacteria. Under the LN regime all lines showed similar classic symptoms of nitrogen deficiency including lower shoot bio-mass and leaf chlorophyll. However, the LN-induced decreases in leaf protein and increases in root protein tended to be smaller in the OCI-1 and OCI-2 lines than in the wild type. When LN-plants were grown with rhizobia, OCI-1 and OCI-2 roots had significantly more crown nodules than wild-type plants. The growth nitrogen regime had a significant effect on the abundance of transcripts encoding vacuolar processing enzymes (VPEs), LN-dependent increases in VPE2 and VPE3 transcripts in all lines. However, the LN-depen-dent increases of VPE2 and VPE3 transcripts were significantly lower in the leaves of OCI-1 and OCI-2 plants than in the wild type. These results show that nitrogen availability regulates the leaf and root cysteine pro-tease, VPE and cystatin transcript profiles in a manner that is in some cases influenced by ectopic OCI expression. Moreover, the OCI-dependent inhibition of papain-like cysteine proteases favours increased nodulation and enhanced tolerance to nitrogen limitation, as shown by the smaller LNdependent decreases in leaf protein observed in the OCI-1 and OCI-2 plants relative to the wild type.

Factors facilitating sustainable scientific partnerships between developed and developing countries

“…Significant benefits come through joint projects on important model plants, such as Arabidopsis, tobacco and poplar, in which the consequences of genetic modification can be tested and the effects of specific gene targets characterized. Such activities result in an increased number and quality of research outputs ( Kunert et al, 2016 ; Quain et al, 2015 ). For example, in total, Professors Kunert and Foyer have published 36 joint peer-reviewed articles in International Journals together.…”

“…Moreover, the associated technologies that were applied in these studies were successfully transferred from Southern Africa to the rest of Africa allowing rapid development in several African partnerships. This partnership also produced improved model and crop plants that have improved tolerance to environmental stresses such as drought and low temperatures ( Kunert et al, 2016 ; Le Roux et al, 2019 ; Quain et al, 2015 ). The development and application of the biotechnological tools for plant improvement against environmental stress has become the major focus of many joint funding applications by scientists worldwide in collaboration with African partners.…”

Search for Nodulation and Nodule Development-Related Cystatin Genes in the Genome of Soybean (Glycine max)

“…Previous studies showed that cystatins could regulate nodulation and nodule development (Quain et al, 2015) and may play a role in regulating proteolysis in the process of nodules senescence and programmed cell death (van Wyk et al, 2014). However, which soybean cystatins play roles in nodulation, nodule development, and senescence is still largely unknown.…”

“…In addition, the inhibitory activities of these cystatins were analyzed and seven of them were actively transcribed in nodules (van Wyk et al, 2014). Ectopic expression of rice phytocystatin positively regulates nodulation and nitrogen deficiency of soybean (Quain et al, 2015). However, almost all of these studies were focused on individual cystatin members.…”

Strigolactones positively regulate chilling tolerance in pea and in Arabidopsis

“…Transgenic tobacco lines ( Nicotiana tabacum ) constitutively expressing OCI showed enhanced tolerance to dark chilling (Van der Vyver et al, ), as well as an increased accumulation of RuBisCo compared to the wild type (Prins et al, ). OCI expression in soybean increased nodule numbers and delayed leaf senescence, as well as leading to a significant increase in drought tolerance (Quain, Makgopa, Cooper, Kunert, & Foyer, ). Phytocystatin expression also prevented stress‐induced programmed cell death in soybean (Solomon, Belenghi, Delledonne, Menachem, & Levine, ).…”