Characterization of two homeodomain transcription factors with critical but distinct roles in virulence in the vascular pathogen Verticillium dahliae

Abstract

Vascular wilt caused by Verticillium dahliae is a destructive disease that represents a chronic economic problem for crop production worldwide. In this work, we characterized two new regulators of pathogenicity in this species. Vph1 (VDAG_06555) was identified in a candidate gene approach as a putative homologue of the transcription factor Ste12. Vhb1 (VDAG_08786), identified in a forward genetics approach, is similar to the homeobox transcription factor Htf1, reported as a regulator of conidiogenesis in several fungi. Deletion of vph1 did not affect vegetative growth, whereas deletion of vhb1 greatly reduced sporulation rates in liquid medium. Both mutants failed to induce Verticillium wilt symptoms. However, unlike Δvph1, Δvhb1 could be re-isolated from the vascular system of some asymptomatic plants. Confocal microscopy further indicated that Δvph1 and Δvhb1 differed in their behaviour in planta; Δvph1 could not penetrate the root cortex, whereas Δvhb1 was impaired in its ability to colonize the xylem. In agreement with these observations, only Δvhb1 could penetrate cellophane paper. On cellophane, wild-type and Δvhb1 strains produced numerous short branches with swollen tips, resembling the hyphopodia formed on root surfaces, contrasting with Δvph1, which generated unbranched long filaments without swollen tips. A microarray analysis showed that these differences in growth were associated with differences in global transcription patterns, and allowed us to identify a large set of novel genes potentially involved in virulence in V. dahliae. Ste12 homologues are known regulators of invasive growth, but Vhb1 is the first putative Htf1 homologue identified with a critical role in virulence.

The Verticillium dahliae Sho1‐MAPK pathway regulates melanin biosynthesis and is required for cotton infection

“…dahliae , several studies have concluded that melanin accumulation is not required for virulence in V . dahliae in hosts such as tobacco, tomato and lettuce (Wang et al ., ; Sarmiento‐Villamil et al ., ; Sarmiento‐Villamil et al ., ), but another study demonstrated that deletion of V . dahliae VdPKS1, a key factor for melanin production, resulted in reduced virulence on cotton (Zhang et al ., ).…”

“…In M. oryzae, melanin is necessary for the strengthening of the appressorium wall prior to host penetration but this phenomenon has not been so clearly demonstrated in other pathogenic fungi producing appressoria, such as Alternaria alternata (Jacobson, 2000). In V. dahliae, several studies have concluded that melanin accumulation is not required for virulence in V. dahliae in hosts such as tobacco, tomato and lettuce (Wang et al, 2018;Sarmiento-Villamil et al, 2018a;Sarmiento-Villamil et al, 2018b), but another study demonstrated that deletion of V. dahliae VdPKS1, a key factor for melanin production, resulted in reduced virulence on cotton . Undoubtedly, gene deletion studies of various MAPK cascade components suggest a strong correlation between melanin biosynthesis and virulence in V. dahliae (Tzima et al, 2010;Tian et al, 2014;Tian et al, 2016;Wang et al, 2016;Yu et al, 2019;Zheng et al, 2019).…”

“…In contrast, analyses of two deletion mutants of either VdPKS1 or VdCmr1 within the melanin biosynthetic gene cluster in V. dahliae strain VdLs.17 revealed that neither was required for full virulence on lettuce and tobacco, but were required for survival in response to UV irradiation and high-temperature stress (Wang et al, 2018). Additionally, melanin accumulation was not necessary for pathogenicity of ΔVst1 strains of V. dahliae, which lack pigment, yet these strains are fully pathogenic on tomato and tobacco (Sarmiento-Villamil et al, 2018a;Sarmiento-Villamil et al, 2018b). Albino mutants or strains, which produce microsclerotia and lack typical pigmentation or that are hyaline, are commonly characterized in V. dahliae, yet are pathogenic (Daayf et al, 1998;Wang et al, 2018).…”

“…Additionally, melanin accumulation was not necessary for pathogenicity of Δ Vst1 strains of V . dahliae , which lack pigment, yet these strains are fully pathogenic on tomato and tobacco (Sarmiento‐Villamil et al ., ; Sarmiento‐Villamil et al ., ). Albino mutants or strains, which produce microsclerotia and lack typical pigmentation or that are hyaline, are commonly characterized in V .…”

“…Host penetration by V. dahliae requires a specialized infection structure called a hyphopodium, which develops following the sensing of extracellular signals from host plants and proper activation of the intracellular signalling pathways via various kinases and other signalling proteins, and is essential for penetration of plant roots Sarmiento-Villamil et al, 2018a;Sarmiento-Villamil et al, 2018b). The surface sensor and signal transduction studies have confirmed the role of these proteins in the pathogenesis in V. dahliae, including the kinases VdMsb (Tian et al, 2014), VdHog1 , VdPbs2 , VdSsk2 and VdSte11 (Yu et al, 2019), VdSsk1 (Zheng et al, 2019) and VMK1 (Tzima et al, 2010).…”