The expression pattern of a rice disease resistance gene xa3/xa26 is differentially regulated by the genetic backgrounds and developmental stages that influence its function

Abstract

Genetic background and developmental stage influence the function of some disease resistance (R) genes. The molecular mechanisms of these modifications remain elusive. Our results show that the two factors are associated with the expression of the R gene in rice Xa3 (also known as Xa26)-mediated resistance to Xanthomonas oryzae pv. oryzae (Xoo), which in turn influences the expression of defense-responsive genes. The background of japonica rice, one of the two major subspecies of Asian cultivated rice, facilitates the function of Xa3 more than the background of indica rice, another rice subspecies. Xa3 expression gradually increases from early seedling stage to adult stage. Japonica plants carrying Xa3 regulated by the native promoter showed an enlarged resistance spectrum (i.e., resistance to more Xoo races), an increased resistance level (i.e., further reduced lesion length), and whole-growth-stage resistance compared to the indica rice; this enhanced resistance was associated with an increased expression of Xa3 throughout the growth stages in the japonica plants, which resulted in enhanced expression of defense-responsive genes. Overexpressing Xa3 with a constitutive strong promoter further enhanced rice resistance due to further increased Xa3 transcripts in both indica and japonica backgrounds, whereas regulating Xa3 with a pathogen-induced weak promoter impaired rice resistance.

Figure 1.—

Performance of Xa3 in different rice lines. Zhonghua 11, Mudanjiang 8, 02428, and Minghui 63 are wild types. Minghui 63 is also the donor of Xa3. The Xa3 gene was driven by native promoter P_Xa3 in plants named with prefix MKbZH, MKb024, or Rb; by P_Ubi in plants named MKbFZH, MKbFMDJ, or MKbFMH; and by P_WRKY13 in plants 12IMKbZH and 12IMKbMDJ. (A) Leaves from transgenic plants and wild types of booting stage at 14 days after inoculation with Xoo strain PXO61. Rb17-2 was a homozygote transgenic line. MKbFMDJ2 and 12IMKbMDJ7 were resistant T₁ plants, and other transgenic plants were T₀ generation. N, negative transgenic plants. (B) Growth of PXO61 in leaves of T₁ transgenic plants at booting stage. The bacterial population was determined from three leaves at each time point by counting colony-forming units (Sun et al. 2004). (C) Growth of PXO61 in leaves of T₁ transgenic plants at the four-leaf stage.

Figure 2.—

Expression level of Xa3 in different rice lines. Each RNA sample was from the mixture of at least three plants. (A) Genetic background influenced Xa3 expression. Transgenic plants named with prefix MKbFMH, MKbFZH, MKbZH, 12IMKbZH, MKbFMDJ, 12IMKbMDJ, and MKb024-1 were resistant T₁ plants, and plants named with prefix Rb were homozygote transgenic lines. All the transgenic plants, except MKbFMH and MKb024-1, in which copy numbers were not determined, carried one copy of Xa3. Xa3 was driven by the native promoter (P_Xa3), maize ubiquitin gene promoter (P_Ubi), or pathogen-induced OsWRKY13 gene promoter (P_WRKY13) in the transgenic plants. (B) Developmental stage influenced Xa3 expression. The expression level of Xa3 in each developmental stage of each rice line was relative to that in the two-leaf stage of Minghui 63.

Figure 3.—

Expression levels of OsWRKY13 and NH1 in different rice lines. Plants were inoculated with Xoo strain PXO61 at booting stage. ck, without inoculation. The expression level of the genes in each time point of each rice line was relative to that in the ck of Minghui 63.

Figure 4.—

Xa3 expression on pathogen infection analyzed by RT–qPCR. Plants were inoculated with Xoo strain PXO61 at booting stage. ck, without inoculation; ckM, transgenic plant MKbZH2 without inoculation.