China is a major producer and consumer of pork. However, the epidemic diseases, such as swine fever and porcine reproductive and respiratory syndrome (Blue-eared Disease, PRRSV), are currently widely spread, which is the greatest difficulty that China's swine industry is facing. The prevention and control of these diseases are thus a major issue to be addressed for the stability and development of swine industry. However, in recent years, the diseases in a swine tend to evolve into multi-pathogen cross-infection, and disease resistance of swine and control effect of a vaccine are significantly reduced. Therefore, for better prevention and control of the diseases, it is required to find new strategies for development of a vaccine or to seek alternative or auxiliary approaches, such as breeding of new varieties of the swine with disease resistance. However, a lot of prior researches have been dedicated to breeding new varieties of livestock with single disease resistance using a specific disease resistance gene, there is thus clearly a large gap for such strategy to satisfy the current reality of frequent occurrence of disease in China's swine industry. Therefore, it becomes the ideal of many researchers that new varieties of the swine with broad-spectrum disease resistance are bred.
Adaptive immune response plays a crucial role in resisting invasion of pathogen by an animal. The activation of T cells is the top priority of the response. Recent studies have shown that effective activation of the T cells requires a two-signal stimulation. One is the signal generated by the binding of the MHC-peptide to TCR, and the other is the second signal generated by the binding of co-stimulatory receptor, primarily CD28, to its ligand. In a normal micro-environment of organism, the antigen-presenting cells often present a small number of antigenic peptides, the binding of which to TCR is not sufficient to activate T cells, thereby causing response impotence of T cells. The co-stimulatory signal pathway provided by T cell co-stimulatory receptors such as CD28 can compensate weaker TCR signaling, thereby activating T cells. Furthermore, when the affinity of TCR with antigen peptides is not sufficient, it is often difficult to activate T cells. The adequate co-stimulatory signals can also play the role of enhancing TCR signaling pathway, so as to activate T cells. In short, sufficient co-stimulatory signals can not only overcome the problem of low rate occupation of TCRs, but also compensate the inadequate TCR affinity. Therefore, co-stimulatory receptor-mediated signaling can enhance the function of adaptive immune system, so that the co-stimulatory receptor becomes a potential candidate for broad-spectrum disease resistance gene.
The gene sequence information and protein function of CD28 receptor, as a typical representative of co-stimulatory receptors, are already described in detail in studies in human and mice. CD28 receptor is recognized as a main co-stimulatory molecule for the initial proliferation and survival of T cells. The receptor can significantly promote activation, proliferation and survival of T cells and can affect the direction of T cell differentiation and up-regulate the expression of cytokines such as IFN-γ when it is linked to its ligand B7.1 (also called CD80) or B7.2 (also called CD86). It has been found that the level of expression of CD28 molecule is significantly down-regulated on the surface of T cells of the aged, and low level of expression of CD28 molecule is also found in some cases of delayed immune system activation. Thus, the expression level of the CD28 molecule directly affects the effective activation of immune system, thereby affecting the disease resistance of the individual organism.
CD28 receptor plays an important role in establishing, enhancing and maintaining T cell immune response, which indicates that CD28 receptor, as a target gene, has good prospects in transgenic breeding research and application. However, the vast majority of relevant studies on co-stimulatory molecules are based on the manner of gene knockout, or removal or enhancement of co-stimulatory signals using anti-CD28 monoclonal antibody. These strategies are obviously unsuitable for breeding research. Furthermore, only predicted genetic sequence information of porcine CD28 receptor is currently published, but its precise coding sequence and protein function haven't been reported so far.