The rust transferred proteins—a new family of effector proteins exhibiting protease inhibitor function

Abstract

Only few fungal effectors have been described to be delivered into the host cell during obligate biotrophic interactions. RTP1p, from the rust fungi Uromyces fabae and U. striatus, was the first fungal protein for which localization within the host cytoplasm could be demonstrated directly. We investigated the occurrence of RTP1 homologues in rust fungi and examined the structural and biochemical characteristics of the corresponding gene products. The analysis of 28 homologues showed that members of the RTP family are most likely to occur ubiquitously in rust fungi and to be specific to the order Pucciniales. Sequence analyses indicated that the structure of the RTPp effectors is bipartite, consisting of a variable N-terminus and a conserved and structured C-terminus. The characterization of Uf-RTP1p mutants showed that four conserved cysteine residues sustain structural stability. Furthermore, the C-terminal domain exhibits similarities to that of cysteine protease inhibitors, and it was shown that Uf-RTP1p and Us-RTP1p are able to inhibit proteolytic activity in Pichia pastoris culture supernatants. We conclude that the RTP1p homologues constitute a rust fungi-specific family of modular effector proteins comprising an unstructured N-terminal domain and a structured C-terminal domain, which exhibit protease inhibitory activity possibly associated with effector function during biotrophic interactions.

The haustorial transcriptomes of Uromyces appendiculatus and Phakopsora pachyrhizi and their candidate effector families

“…These proteins represent 3.5% of the haustorial transcriptome, and 40 (26%) of these had GO annotations. These putative secreted proteins include homologues to haustoria‐expressed secreted proteins (HESP379, HESP767, HESP‐C49; Catanzariti et al ., ) from flax rust, and to rust transferred proteins (RTPs) in Uromyces species (Pretsch et al ., ).…”

“…The secreted proteins include expansin homologues, one protein with a small secreted protein (SSP) annotation and three RTPs. Two have been identified previously, Ua‐ RTP1p and Ua‐ RTP2p (Puthoff et al ., ), but the third is more distantly related and can be classified as RTP9p‐like (Pretsch et al ., ). The 15 reads for Ua_contig00666 suggest that Ua‐RTP9 is weakly expressed relative to Ua‐RTP1 (Ua_contig06414, 355 reads) and Ua‐RTP2 (Ua_contig06302, 296 reads).…”

Expression profiling and functional analyses of BghPTR2, a peptide transporter from Blumeria graminis f. sp. hordei

“…In Colletotrichum, a hemibiotroph, expression of nutrient transporters were predominantly found in the transition from biotrophy to necrotrophy and the haustorial compartment is hypothesized to be mainly responsible for delivery of effector molecules and secondary metabolites to influence plant response processes (O'Connell et al 2012). Pretsch et al (2013) reports that haustorium specific effector proteins from a range of rust fungi exhibit protease inhibitory activity which could protect effector proteins from degradation in the biotrophic interaction. This implies that PTR2s would have limited substrates for transport while the uptake of glucose at the haustorial interface reported by Mendgen & Nass (1988) would be unaffected by this.…”

Secreted biomolecules in fungal plant pathogenesis

“…The initial functional characterization of putative secreted fungal effectors is now taking place through their in planta heterologous over-expression using a new endophytic bacterial delivery system (Upadhyaya et al, 2014). In non-cereal infecting rusts, Uromyces fabae and Uromyces striatus, the rust transferred protein 1 (RTP1p) accumulates around the haustorial complex, forming filamentous structures in the EHM that are predicted to stabilize the host cell and protect the haustorium from degrada tion Pretsch et al, 2013). The homologous RTP1 gene in P. striiformis f. sp.…”