Increased Transcriptions and Pathogenicity

Since extensive description of P. brasiliensis yeast-expressed and phase-transition related genes can be found in the various reports listed above, this section discusses representative genes that seem to be experimentally related to the fungal interaction with the host.

So far the best approach to studying P. brasiliensis gene transcriptional response to host cell contact is the one by Tavares et al. (2005) who studied regulation of 1,152 selected fungal genes after 6 h of coculture with peritoneal murine macrophages, with 90% of yeast cells internalized or adherent to phagocytes. Microarray hybridization revealed differential expression of 152 Pb01 transcripts considering a twofold variation in comparison with transcription during in vitro growth. Apparently, P. brasiliensis reacted to macrophage nitrogen and oxygen radicals by increasing transcription of genes related to oxidative stress. In this sense, SOD3 (Cu-Zn superoxide dismutase), which encodes a predicted GPI-anchored plasma membrane protein (Castro et al., 2005), was highly expressed, and HSP60 and QCR8 (subunit VIII of citocrome oxidase c) mRNAs were also above the twofold cutoff. Histone-related and ribosomal subunit genes were also upregulated, while the P-1,3-glucan synthase gene was downregulated. It should be pointed out that the a-1,3-glucan synthase gene is highly expressed in yeast cells (Marques et al., 2004).

The most abundant transcripts detected by Bailao et al. (2006) in yeasts recovered from liver were HSP30 and ZTR1 (high-affinity zinc/iron permease), followed by CRT3 (high-affinity copper transporter) and GADPH (glyceraldehyde 3-phosphate dehydrogenase). While ZTR1 and CRT3 transcripts might have been upregulated in response to iron limitation in the organ, GADPH as discussed above adheres to ECM-associated proteins (Barbosa et al., 2004), and may contribute to fungal invasion. Two serine proteinase gene homologs more expressed in host conditions (PR1H and SP1) may contribute to the process by cleaving local proteins.

It should be pointed out that the glyoxylate cycle-specific genes ICL1 and MLS1, encoding isocitrate lyase and malate synthase, have not been differentially detected in P. brasiliensis cultivated with macrophages or human blood, in contrast to C. albicans (Fradin et al., 2003). These enzymes are C. albicans virulence factors and potential drug targets due to their absence in humans (Lorenz et al., 2001).

During temperature-driven mycelium-to-yeast transition of Pb18 in rich medium, a total of 2,583 genes from a 4,692-element microarray were modulated, showing the complexity of a phenomenon that involves amino acid catabolism, signal transduction, protein synthesis, cell wall metabolism, genome structure, heat and oxidative stress response, growth control, and development (Nunes et al., 2005). The most interesting gene clusters involved transcripts whose levels increased sharply after either 5 or 48 h of temperature shift and remained high until 120 h, when over 80% of the cells were transformed into yeasts. Together, these two clusters involved 30 genes, among which there were two related to amino acid catabolism, encoding a homolog of 4-hydroxyphenyl-pyruvate dehydrogenase (4-HPPD), the C. immitis protective T-cell stimulatory antigen, and a branched chain a-keto acid dehydrogenase. The relevance of amino acid catabolism to mycelium-to-yeast conversion and yeast-phase maintenance was determined by the specific inhibitory effect of nitisinone in the dimorphic transition to yeast and yeast growth. Nitisinone, or NTBC (2-(2-nitro-4-trifluoromethylbenzoyl)-cyclohexane-1,3-dione), is a commercially available inhibitor of 4-HPPD used in treatment of hereditary type I tyrosinemia, and could be a completely new alternative in the treatment of PCM. More importantly, Nunes et al. (2005) tested a series of new NTBC derivatives and found that compound 8 is 40 times more potent than nitisinone in the inhibition of yeast cells development.

Included in the two clusters mentioned above (Nunes et al., 2005) are also PbGP43, METR, encoding a zinc-finger transcription factor related to induction of genes of the sulfate assimilation pathway (Ferreira et al., 2006), PbY20, and gene homologs encoding an alcohol dehydrogenase III and a formamidase. The for-mamidase gene (PbFMD) has recently been characterized (Borges et al., 2005) and the antigenic properties of the gene product were mentioned in a previous section. Upon contact with macrophages, however, PbFMD was downregulated. The PbY20 gene has recently been characterized (Daher et al., 2005). It encodes a member of the flavodoxin-like WrbA family of quinone reductases, which might be involved in protection against oxygen intermediates. PbY20 protein, together with enolase, has originally been recognized as yeast-preferential by comparing 2-D profiles of total proteins extracted from fully formed mycelium and yeast cells (Cunha et al., 1999). The transcript and the protein are poorly expressed in mycelium cells (Daher et al., 2005), which start to overexpress Pb Y20 mRNA only after 48 h of temperature shift for transition to yeast cells that express the transcript in large amounts (Felipe et al., 2005; Nunes et al., 2005). The protein was mainly localized to large cytoplasmic vacuoles and in the cell wall. PbY20 might act as an antioxidant in vivo, but this function remains to be proved. A 61 kDa catalase, possibly peroxisomal, has been detected in yeast fungal extracts with PCM patients' sera (Fonseca et al., 2001; Moreira et al., 2004). Both transcript and protein were upregulated during transition to yeast and an increase in protein levels was already apparent 20 min after H2O2 addition, suggesting that it has an antioxidant role. We have recently characterized two adjacent genes, PbLON and PbMDJ1, encoding mitochondrially sorted ATP-dependent proteinase and an Hsp40, respectively (Barros et al., 2001; Batista et al.,

2006a). The genes share a promoter region that bears 3 mapped heat-shock elements and one AP-1-binding motif directed to PbLON, which upregulates fivefold upon addition of H2O2 (Batista et al., 2006b).

Cure Your Yeast Infection For Good

Cure Your Yeast Infection For Good

The term vaginitis is one that is applied to any inflammation or infection of the vagina, and there are many different conditions that are categorized together under this ‘broad’ heading, including bacterial vaginosis, trichomoniasis and non-infectious vaginitis.

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