Non-coding RNAs associated with IS200/605: Identification and functional characterizal in the archaea Halobacterium salinarum NRC-1

Author(s)

José Vicente Gomes Filho

• Document type: Doctoral Thesis
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2017-06-13

Examining board members

Tie Koide; Angela Kaysel Cruz; José Freire da Silva Neto; Alessandro de Mello Varani

• Advisor: Tie Koide

Abstract

Mobile genetic elements (MGEs) are extremely important for plasticity and evolution of genomes. Their impacts are diverse and could be related to antibiotic resistence or symbiosis. One of the most important class of MGEs are insertion sequences (ISs). These elements are found widespread throughout bacteria and archaea, presenting a great diversity of families. An interesting family is the IS200/605, this family is found widespread in bacteria, archaea and viruses and is divided in three subgroups according to it\'s genetic composition: IS200 with tnpA gene alone, IS605 with tnpA and tnpB and IS1341 with tnpB only. Another interesting aspect is the utilization of single-stranded DNA as a substrate during the transposition process. In this work, through the analysis of public available transcriptomic data we identified transcripts that overlaps the 3\' end of tnpB in IS200/605 in both bacteria and archaea. These transcripts were named sense overlapping transcripts (sotRNAs). Sequence and secondary structure analysis showed a conserved motif present in sotRNAs, the RE-like motif. Using the consensus sequence of this motif we identified novel intergenic ncRNAs containing this motif that are derived from IS200/605. For functional characterization we overexpressed a sotRNA (VNG_sot0042) and a intergenic (VNG_R0052), both containing the RE-like motif. Standard growth curves demonstrated that the overexpression of these ncRNAs improve H. salinarum growth showing that this RNA is functional. To further evaluate the impact of the overexpressions we prepared RNA-seq libraries of the strain overexpressing VNG_sot0042 and in parallel performed SILAC experiments to identify potential protein-RNA interaction partners. Differentially regulated genes and interacting proteins associated with adhesion and persistent cells generation were found. Adhesion and survival assays showed that the lineage overexpressing VNG_sot0042 has a better capability to adhere in glass surfaces and survive more in diverse stressful conditions. (AU)

• FAPESP's process: 13/21522-5 - Functional characterization of non-coding RNAs associated with IS1341 transposons in Halobacterium salinarum
• Grantee: José Vicente Gomes Filho
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)