A Bacillus licheniformis strain, I89, isolated from a hot spring environment in the Azores, Portu... more A Bacillus licheniformis strain, I89, isolated from a hot spring environment in the Azores, Portugal, strongly inhibited growth of Gram-positive bacteria. It produced a peptide antibiotic at 50 °C. The antibiotic was purified and biochemically characterized. It was highly resistant to several proteolytic enzymes. Additionally, it retained its antimicrobial activity after incubation at pH values between 3.5 and 8; it was thermostable, retaining about 85% and 20% of its activity after 6 h at 50 °C and 100 °C, respectively. Its molecular mass determined by mass spectrometry was 3249.7 Da.
Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a CTG expansion in the 3' U... more Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a CTG expansion in the 3' UTR of the DMPK gene. It has been hypothesized that the pathogenesis in DM1 is triggered by a toxic gain of function of the expanded DMPK RNA. This expanded RNA is retained in nuclear foci where it sequesters and induces alterations in the levels of RNA-binding proteins. To model DM1 and study the implication of RNA-binding proteins in CUG-induced toxicity, we have generated a Drosophila DM1 model expressing a non-coding mRNA containing 480 interrupted CUG repeats; i.e., [(CUG) 20 CUCGA] 24 . This (iCUG) 480 transcript accumulates in nuclear foci and its expression leads to muscle wasting and degeneration in Drosophila. We also report that altering the levels of two RNA-binding proteins known to be involved in DM1 pathogenesis, MBNL1 and CUGBP1, modify the (iCUG) 480 degenerative phenotypes.
ETR-3 (also know as BRUNOL3, NAPOR, and CUGBP2) is one of six members of the CELF (CUG-BP1-and ET... more ETR-3 (also know as BRUNOL3, NAPOR, and CUGBP2) is one of six members of the CELF (CUG-BP1-and ETR-3-like factor) family of splicing regulators. ETR-3 regulates splicing by direct binding to the pre-mRNA. We performed systematic evolution of ligands by exponential enrichment (SELEX) to identify the preferred binding sequence of ETR-3. After five rounds of SELEX, ETR-3 selected UG-rich sequences, in particular UG repeats and UGUU motifs. Either of these selected motifs was able to restore ETR-3 binding and responsiveness to a nonresponsive splicing reporter in vivo. Moreover, this effect was not specific to ETR-3 since minigenes containing either of the two motifs were responsive to two other CELF proteins (CUG-BP1 and CELF4), indicating that different members of the CELF family can mediate their effects via a common binding site. Using the SELEX-identified motifs to search the human genome, we identified several possible new ETR-3 targets. We created minigenes for two of these genes, the CFTR and MTMR1 genes, and confirmed that ETR-3 regulates their splicing patterns. For the CFTR minigene this regulation was demonstrated to be dependent on the presence of the putative binding site identified in our screen. These results validate this approach to search for new targets for RNA processing proteins.
A Bacillus licheniformis strain, I89, isolated from a hot spring environment in the Azores, Portu... more A Bacillus licheniformis strain, I89, isolated from a hot spring environment in the Azores, Portugal, strongly inhibited growth of Gram-positive bacteria. It produced a peptide antibiotic at 50 °C. The antibiotic was purified and biochemically characterized. It was highly resistant to several proteolytic enzymes. Additionally, it retained its antimicrobial activity after incubation at pH values between 3.5 and 8; it was thermostable, retaining about 85% and 20% of its activity after 6 h at 50 °C and 100 °C, respectively. Its molecular mass determined by mass spectrometry was 3249.7 Da.
Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a CTG expansion in the 3' U... more Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a CTG expansion in the 3' UTR of the DMPK gene. It has been hypothesized that the pathogenesis in DM1 is triggered by a toxic gain of function of the expanded DMPK RNA. This expanded RNA is retained in nuclear foci where it sequesters and induces alterations in the levels of RNA-binding proteins. To model DM1 and study the implication of RNA-binding proteins in CUG-induced toxicity, we have generated a Drosophila DM1 model expressing a non-coding mRNA containing 480 interrupted CUG repeats; i.e., [(CUG) 20 CUCGA] 24 . This (iCUG) 480 transcript accumulates in nuclear foci and its expression leads to muscle wasting and degeneration in Drosophila. We also report that altering the levels of two RNA-binding proteins known to be involved in DM1 pathogenesis, MBNL1 and CUGBP1, modify the (iCUG) 480 degenerative phenotypes.
ETR-3 (also know as BRUNOL3, NAPOR, and CUGBP2) is one of six members of the CELF (CUG-BP1-and ET... more ETR-3 (also know as BRUNOL3, NAPOR, and CUGBP2) is one of six members of the CELF (CUG-BP1-and ETR-3-like factor) family of splicing regulators. ETR-3 regulates splicing by direct binding to the pre-mRNA. We performed systematic evolution of ligands by exponential enrichment (SELEX) to identify the preferred binding sequence of ETR-3. After five rounds of SELEX, ETR-3 selected UG-rich sequences, in particular UG repeats and UGUU motifs. Either of these selected motifs was able to restore ETR-3 binding and responsiveness to a nonresponsive splicing reporter in vivo. Moreover, this effect was not specific to ETR-3 since minigenes containing either of the two motifs were responsive to two other CELF proteins (CUG-BP1 and CELF4), indicating that different members of the CELF family can mediate their effects via a common binding site. Using the SELEX-identified motifs to search the human genome, we identified several possible new ETR-3 targets. We created minigenes for two of these genes, the CFTR and MTMR1 genes, and confirmed that ETR-3 regulates their splicing patterns. For the CFTR minigene this regulation was demonstrated to be dependent on the presence of the putative binding site identified in our screen. These results validate this approach to search for new targets for RNA processing proteins.
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Papers by nuno faustino