Extremophilic Acinetobacter strains from High-Altitude Lakes in Argentinean Puna: UV-B resistance and DNA repairing mechanisms involving photolyases.

Abstract

High-Altitude Andean Lakes (HAAL) at the northwest of Argentina (from 3,000 to 6,000 masl) are extreme environments of biotechnological interest. A collection of 200 extremophilic strains which displayed multiple resistance profiles to diverse environmental stress: hipersalinity, high UV-B irradiation and arsenic. has been built. The aim of this work is to study the UV-resistance profile of four extremophilic strains from the genus Acinetobacter by partially characterize their photorepairing mechanisms upon UV-B DNA damage. In addition, we present Acinetobacter sp. Ver3 whose recent genome sequencing revealed the presence of interesting and novel photoreceptors such as photolyases.Acinetobacter by partially characterize their photorepairing mechanisms upon UV-B DNA damage. In addition, we present Acinetobacter sp. Ver3 whose recent genome sequencing revealed the presence of interesting and novel photoreceptors such as photolyases.Acinetobacter sp. Ver3 whose recent genome sequencing revealed the presence of interesting and novel photoreceptors such as photolyases. Acinetobacter spp. N40, Ver3, Ver5, Ver7 and sensitive strains Acinetobacter baumannii DSM 30007 and Acinetobacter jhonsonnii DSM 6963 where exposed to UV-B light (38 kJ) in liquid media. The control strains were not able to maintain their population after the UV-B exposure while Acinetobacter sp. Ver7 showed 100% survival after the irradiation, Acinetobacter sp. Ver5, 50% and Acinetobacter sp. Ver3 and N40 20% and 1% respectively. After the DNA damage, the strains were exposed to light (PR) and dark repair (DR) during 2 h and DNA photoproducts in total genomic DNA after each treatment was measured using HPLC-MS/MS. Acinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. Acinetobacterspp. N40, Ver3, Ver5, Ver7 and sensitive strains Acinetobacter baumannii DSM 30007 and Acinetobacter jhonsonnii DSM 6963 where exposed to UV-B light (38 kJ) in liquid media. The control strains were not able to maintain their population after the UV-B exposure while Acinetobacter sp. Ver7 showed 100% survival after the irradiation, Acinetobacter sp. Ver5, 50% and Acinetobacter sp. Ver3 and N40 20% and 1% respectively. After the DNA damage, the strains were exposed to light (PR) and dark repair (DR) during 2 h and DNA photoproducts in total genomic DNA after each treatment was measured using HPLC-MS/MS. Acinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. AcinetobacterAcinetobacter jhonsonnii DSM 6963 where exposed to UV-B light (38 kJ) in liquid media. The control strains were not able to maintain their population after the UV-B exposure while Acinetobacter sp. Ver7 showed 100% survival after the irradiation, Acinetobacter sp. Ver5, 50% and Acinetobacter sp. Ver3 and N40 20% and 1% respectively. After the DNA damage, the strains were exposed to light (PR) and dark repair (DR) during 2 h and DNA photoproducts in total genomic DNA after each treatment was measured using HPLC-MS/MS. Acinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. AcinetobacterAcinetobacter sp. Ver7 showed 100% survival after the irradiation, Acinetobacter sp. Ver5, 50% and Acinetobacter sp. Ver3 and N40 20% and 1% respectively. After the DNA damage, the strains were exposed to light (PR) and dark repair (DR) during 2 h and DNA photoproducts in total genomic DNA after each treatment was measured using HPLC-MS/MS. Acinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. AcinetobacterAcinetobacter sp. Ver3 and N40 20% and 1% respectively. After the DNA damage, the strains were exposed to light (PR) and dark repair (DR) during 2 h and DNA photoproducts in total genomic DNA after each treatment was measured using HPLC-MS/MS. Acinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. AcinetobacterAcinetobacter sp. Ver3 and Ver5 were able to completely recover their initial population after PR. AcinetobacterAcinetobacter sp. N40 also was able to increase their population thanks to the PR (60%). In contrast, A. baummanii was not able to recover after PR, while A. jhonsonnii recovered partially in a 15%. DR was not efficient for recovering the initial population for all strains. The most efficient strains to deplete TT-CPD and CT-CPD under PR were Acinetobacter sp. Ver3 and Ver7 with a reduction of 90% and 100% compared to the controls non-PR, respectively. Genome analyses (performed after pyro-sequencing and RAST annotation) revealed the presence of two different photolyase-coding sequences in Ver3. Upon BLAST analysis the nearest matches of one of them (PL1) was found to be the photolyase (ZP06727183) from A. haemolyticus ATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsoniiA. baummanii was not able to recover after PR, while A. jhonsonnii recovered partially in a 15%. DR was not efficient for recovering the initial population for all strains. The most efficient strains to deplete TT-CPD and CT-CPD under PR were Acinetobacter sp. Ver3 and Ver7 with a reduction of 90% and 100% compared to the controls non-PR, respectively. Genome analyses (performed after pyro-sequencing and RAST annotation) revealed the presence of two different photolyase-coding sequences in Ver3. Upon BLAST analysis the nearest matches of one of them (PL1) was found to be the photolyase (ZP06727183) from A. haemolyticus ATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsoniiwas not able to recover after PR, while A. jhonsonnii recovered partially in a 15%. DR was not efficient for recovering the initial population for all strains. The most efficient strains to deplete TT-CPD and CT-CPD under PR were Acinetobacter sp. Ver3 and Ver7 with a reduction of 90% and 100% compared to the controls non-PR, respectively. Genome analyses (performed after pyro-sequencing and RAST annotation) revealed the presence of two different photolyase-coding sequences in Ver3. Upon BLAST analysis the nearest matches of one of them (PL1) was found to be the photolyase (ZP06727183) from A. haemolyticus ATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsoniiAcinetobacter sp. Ver3 and Ver7 with a reduction of 90% and 100% compared to the controls non-PR, respectively. Genome analyses (performed after pyro-sequencing and RAST annotation) revealed the presence of two different photolyase-coding sequences in Ver3. Upon BLAST analysis the nearest matches of one of them (PL1) was found to be the photolyase (ZP06727183) from A. haemolyticus ATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsoniiA. haemolyticus ATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsoniiATCC 19194 (62% identity) and the photolyase (ZP06062937) from A. johnsonii SH046 (61% identity). In turn, the nearest matches for the second photolyase (PL2) were the deoxyribodipyrimidine photolyase-related protein (ZP06693260) from Acinetobacter sp. SH024 (70% identity) and the photolyase (ZP06066462) from A. junii SH205 (61% identity). The conserved domain architecture retrieval tool from the NCBI database revealed a different domain structure for the studied photolyases. Based on homology modeling, a strong three-dimensional similarity to the photolyase from E. coli (PDB 1DNPA) was observed for the PL1. Overexpression of the PL1-coding sequence revealed it can be functional and improve UV-B resistance profiles in E. coli cells. Functional characterization of the purified photolyase proteins is ongoing.Acinetobacter sp. SH024 (70% identity) and the photolyase (ZP06066462) from A. junii SH205 (61% identity). The conserved domain architecture retrieval tool from the NCBI database revealed a different domain structure for the studied photolyases. Based on homology modeling, a strong three-dimensional similarity to the photolyase from E. coli (PDB 1DNPA) was observed for the PL1. Overexpression of the PL1-coding sequence revealed it can be functional and improve UV-B resistance profiles in E. coli cells. Functional characterization of the purified photolyase proteins is ongoing.A. junii SH205 (61% identity). The conserved domain architecture retrieval tool from the NCBI database revealed a different domain structure for the studied photolyases. Based on homology modeling, a strong three-dimensional similarity to the photolyase from E. coli (PDB 1DNPA) was observed for the PL1. Overexpression of the PL1-coding sequence revealed it can be functional and improve UV-B resistance profiles in E. coli cells. Functional characterization of the purified photolyase proteins is ongoing.E. coli (PDB 1DNPA) was observed for the PL1. Overexpression of the PL1-coding sequence revealed it can be functional and improve UV-B resistance profiles in E. coli cells. Functional characterization of the purified photolyase proteins is ongoing.E. coli cells. Functional characterization of the purified photolyase proteins is ongoing.