g. the Trehalose Phosphorylase pathway, for which putative genes have been identified and partially characterized in N. crassa and A. fumigatus and also exist in A. niger (ANI_1_2720024). However, it is possible to generate mutants,
within the homologous Tps/Tpp group, in A. fumigatus and A. nidulans that totally lack trehalose [11, 12]. Therefore, we believe that this is the only active trehalose synthesis pathway in Aspergilli. However, internal trehalose contents may not solely be dependent on the presence and expression of these six genes, as in S. cerevisiae there is a strong linkage between trehalose synthesis and the degrading trehalases  as well as evidences of posttranscriptional activation of the genes involved in trehalose metabolism Selleck CP673451 [42, 43]. Besides a putative phosphatase activity, TppB and TppC may have similar biological roles as the yeast proteins Tps3 and Tsl1, which also contain phosphatase domains – in yeasts, deletion of both genes is necessary before some reduction in internal trehalose content can be observed . It is intriguing that tpsB and tppC are linked on the chromosome. We cannot explain why the conidial trehalose content in this double mutant was significantly higher
after 28 days, but based on the expression AZD5582 supplier patterns (see Figure 3), it is possible that the expression of the two genes are regulated by the same factors. In addition to the above-mentioned observations, some conclusions can be drawn from the gene expression data: All identified genes were expressed, indicating that the paralogs are not inactive duplicates. For tpsC and tppB, the expressions were consistently low after 6 h, indicating that the two genes may be regulated by the same mechanism. This assumption is supported by a previous observation using A. oryzae arrays where the tpsC and tppB orthologs were down-regulated in a deletion strain of atfA,
a gene encoding a transcription factor . To our knowledge, two previous studies describing the expression of LY294002 trehalose synthesis genes in A. niger during germination, using microarray technology, or in combination with RNA sequencing, have been published [29, 45]. With the exception that van Leeuwen and co-workers  saw a drastic drop after 2 h and then a gradual up-regulation of tpsA and tpsB, those results are in line with our findings. The extensive measurements of internal trehalose indicate that the trehalose contents, for all strains, were low in 5 day old conidia, significantly elevated in 14 day old conidia, and then maintained at the value of 14 days (Figure 7). A plausible hypothesis is that conidia of A. niger reach full maturity, at least in terms of trehalose accumulation, sometime between 5 days and 2 weeks.