Analysis of Expressed Sequence Tags fromUromyces appendiculatusHyphae and Haustoria and Their Comparison to Sequences from Other Rust Fungi

Abstract

Hyphae, 2 to 8 days postinoculation (dpi), and haustoria, 5 dpi, were isolated from Uromyces appendiculatus infected bean leaves (Phaseolus vulgaris cv. Pinto 111) and a separate cDNA library prepared for each fungal preparation. Approximately 10,000 hyphae and 2,700 haustoria clones were sequenced from both the 5' and 3' ends. Assembly of all of the fungal sequences yielded 3,359 contigs and 927 singletons. The U. appendiculatus sequences were compared with sequence data for other rust fungi, Phakopsora pachyrhizi, Uromyces fabae, and Puccinia graminis. The U. appendiculatus haustoria library included a large number of genes with unknown cellular function; however, summation of sequences of known cellular function suggested that haustoria at 5 dpi had fewer transcripts linked to protein synthesis in favor of energy metabolism and nutrient uptake. In addition, open reading frames in the U. appendiculatus data set with an N-terminal signal peptide were identified and compared with other proteins putatively secreted from rust fungi. In this regard, a small family of putatively secreted RTP1-like proteins was identified in U. appendiculatus and P. graminis.

Expression profiling of genes involved in the biotrophic colonisation of Coffea arabica leaves by Hemileia vastatrix

“…In H. vastatrix, RTP1 transcripts were highly expressed at all in planta stages and for all compatible interactions surveyed, but not in the in vitro phases. These results are consistent with the expression profiles of orthologous RTP1 genes in M. larici-populina (Duplessis et al 2011b), P. triticina (Song et al 2011) and U. appendiculatus (Puthoff et al 2008). Several RTP gene homologs were detected in a 454-pyrosequencing library of rust infected coffee leaves , indicating their expression in planta.…”

“…Besides the CDs and AATs, for which the occurrence of genes families complicated resolution of orthology with other rust fungi, H. vastatrix RTP1 and MAPK genes did not show lower similarities in comparison to other rust fungi. For RTP1, this result is likely the consequence of the low similarity among rust fungi and the presence of a short and highly conserved central region (as detailed in the Online Resource; Puthoff et al 2008;Fernandez et al 2011). On the contrary, MAPK genes are highly conserved between all rust species, which could indicate a purifying selection pressure, thus a pivotal role in signalling during host infection.…”

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

“…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).…”

Proteome analysis of wheat leaf rust fungus, Puccinia triticina, infection structures enriched for haustoria

“…This gene was subsequently shown to be expressed exclusively in haustoria and its protein product was transferred from the extra‐haustorial matrix to the host nucleus 42. Its homolog from Uromyces appendiculatus , Ua‐RTP1 , has been used to demonstrate haustorium‐specific enrichment of fungal RNA over plant RNA 43, 44. We identified a single homolog in Pt , PtRTP1 , and measured by qRT‐PCR its expression in resting urediniospores, urediniospores germinating in vitro over water, in the infected wheat leaf starting material and in the haustorial‐enriched fractions.…”