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Title: Identifying Aspects of the Post-Transcriptional Program Governing the Proteome of the Green Alga Micromonas pusilla

Abstract

Micromonas is a unicellular green alga that belongs to the prasinophytes, a sister lineage to land plants. This picoeukaryotic (<2 μm diameter) alga is widespread in the marine environment but still not understood at the cellular level. Here, we examine the mRNA and protein level changes that take place over the course of the day-night cycle using mid-exponential nutrient replete cultures of Micromonas pusilla CCMP1545 grown and analyzed in biological triplicate. During the experiment, samples were collected at key transition points during the diel for evaluation using high-throughput LC-MS proteomics. We also sequenced matched mRNA samples from the same time points, using pair-ended directional Illumina RNA-Seq to investigate the dynamics and relationship between the mRNA and protein expression programs of M. pusilla. Similar to a prior study of the marine cyanobacterium Prochlorococcus, we found significant divergence in the mRNA and proteomics expression dynamics in response to the light:dark cycle. Additionally, expressional responses of genes and the proteins they encoded could also be variable within the same metabolic pathway, such as the oxygenic photosynthesis pathway. A regression framework was used to predict protein levels using both mRNA expression and gene-specific sequence-based features. Several features in the genome sequence were found tomore » influence protein abundance including the codon usage and the length of the 3’ UTR. Collectively, our studies provide insights into the regulation of the proteome over a diel as relationships between the transcriptional and translational programs in the widespread marine green alga Micromonas.« less

Authors:
; ; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1342328
Report Number(s):
PNNL-SA-115265
Journal ID: ISSN 1932-6203; 47672; KP1601030
DOE Contract Number:
AC05-76RL01830
Resource Type:
Journal Article
Resource Relation:
Journal Name: PLoS ONE; Journal Volume: 11; Journal Issue: 7
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; Environmental Molecular Sciences Laboratory

Citation Formats

Waltman, Peter H., Guo, Jian, Reistetter, Emily Nahas, Purvine, Samuel, Ansong, Charles K., van Baren, Marijke J., Wong, Chee-Hong, Wei, Chia-Lin, Smith, Richard D., Callister, Stephen J., Stuart, Joshua M., Worden, Alexandra Z., and Mills, Ken. Identifying Aspects of the Post-Transcriptional Program Governing the Proteome of the Green Alga Micromonas pusilla. United States: N. p., 2016. Web. doi:10.1371/journal.pone.0155839.
Waltman, Peter H., Guo, Jian, Reistetter, Emily Nahas, Purvine, Samuel, Ansong, Charles K., van Baren, Marijke J., Wong, Chee-Hong, Wei, Chia-Lin, Smith, Richard D., Callister, Stephen J., Stuart, Joshua M., Worden, Alexandra Z., & Mills, Ken. Identifying Aspects of the Post-Transcriptional Program Governing the Proteome of the Green Alga Micromonas pusilla. United States. doi:10.1371/journal.pone.0155839.
Waltman, Peter H., Guo, Jian, Reistetter, Emily Nahas, Purvine, Samuel, Ansong, Charles K., van Baren, Marijke J., Wong, Chee-Hong, Wei, Chia-Lin, Smith, Richard D., Callister, Stephen J., Stuart, Joshua M., Worden, Alexandra Z., and Mills, Ken. 2016. "Identifying Aspects of the Post-Transcriptional Program Governing the Proteome of the Green Alga Micromonas pusilla". United States. doi:10.1371/journal.pone.0155839.
@article{osti_1342328,
title = {Identifying Aspects of the Post-Transcriptional Program Governing the Proteome of the Green Alga Micromonas pusilla},
author = {Waltman, Peter H. and Guo, Jian and Reistetter, Emily Nahas and Purvine, Samuel and Ansong, Charles K. and van Baren, Marijke J. and Wong, Chee-Hong and Wei, Chia-Lin and Smith, Richard D. and Callister, Stephen J. and Stuart, Joshua M. and Worden, Alexandra Z. and Mills, Ken},
abstractNote = {Micromonas is a unicellular green alga that belongs to the prasinophytes, a sister lineage to land plants. This picoeukaryotic (<2 μm diameter) alga is widespread in the marine environment but still not understood at the cellular level. Here, we examine the mRNA and protein level changes that take place over the course of the day-night cycle using mid-exponential nutrient replete cultures of Micromonas pusilla CCMP1545 grown and analyzed in biological triplicate. During the experiment, samples were collected at key transition points during the diel for evaluation using high-throughput LC-MS proteomics. We also sequenced matched mRNA samples from the same time points, using pair-ended directional Illumina RNA-Seq to investigate the dynamics and relationship between the mRNA and protein expression programs of M. pusilla. Similar to a prior study of the marine cyanobacterium Prochlorococcus, we found significant divergence in the mRNA and proteomics expression dynamics in response to the light:dark cycle. Additionally, expressional responses of genes and the proteins they encoded could also be variable within the same metabolic pathway, such as the oxygenic photosynthesis pathway. A regression framework was used to predict protein levels using both mRNA expression and gene-specific sequence-based features. Several features in the genome sequence were found to influence protein abundance including the codon usage and the length of the 3’ UTR. Collectively, our studies provide insights into the regulation of the proteome over a diel as relationships between the transcriptional and translational programs in the widespread marine green alga Micromonas.},
doi = {10.1371/journal.pone.0155839},
journal = {PLoS ONE},
number = 7,
volume = 11,
place = {United States},
year = 2016,
month = 7
}
  • The surface waters of oligotrophic oceans have chronically low phosphate (P i) concentrations, which renders dissolved organic phosphorus (DOP) an important nutrient source. In the subtropical North Atlantic, cyanobacteria are often numerically dominant, but picoeukaryotes can dominate autotrophic biomass and productivity making them important contributors to the ocean carbon cycle. Despite their importance, little is known regarding the metabolic response of picoeukaryotes to changes in phosphorus (P) source and availability. To understand the molecular mechanisms that regulate P utilization in oligotrophic environments, we evaluated transcriptomes of the picoeukaryote Micromonas pusilla grown under P i-replete and -deficient conditions, with an additionalmore » investigation of growth on DOP in replete conditions. Genes that function in sulfolipid substitution and P i uptake increased in expression with P i-deficiency, suggesting cells were reallocating cellular P and increasing P acquisition capabilities. P i-deficient M. pusilla cells also increased alkaline phosphatase activity and reduced their cellular P content. Cells grown with DOP were able to maintain relatively high growth rates, however the transcriptomic response was more similar to the P i-deficient response than that seen in cells grown under P i-replete conditions. The results demonstrate that not all P sources are the same for growth; while M. pusilla, a model picoeukaryote, may grow well on DOP, the metabolic demand is greater than growth on P i. Lastly, these findings provide insight into the cellular strategies which may be used to support growth in a stratified future ocean predicted to favor picoeukaryotes.« less
  • Picoeukaryotes are a taxonomically diverse group of organisms less than 2 micrometers in diameter. Photosynthetic marine picoeukaryotes in the genus Micromonas thrive in ecosystems ranging from tropical to polar and could serve as sentinel organisms for biogeochemical fluxes of modern oceans during climate change. These broadly distributed primary producers belong to an anciently diverged sister clade to land plants. Although Micromonas isolates have high 18S ribosomal RNA gene identity, we found that genomes from two isolates shared only 90percent of their predicted genes. Their independent evolutionary paths were emphasized by distinct riboswitch arrangements as well as the discovery of intronicmore » repeat elements in one isolate, and in metagenomic data, but not in other genomes. Divergence appears to have been facilitated by selection and acquisition processes that actively shape the repertoire of genes that are mutually exclusive between the two isolates differently than the core genes. Analyses of the Micromonas genomes offer valuable insights into ecological differentiation and the dynamic nature of early plant evolution.« less
  • The phosphorus-limited growth kinetics of the chlorophyte Scenedesmus quadricauda and the cyanophyte Synechococcus Nageli were studied by using batch and continuous culturing techniques. The steady-state phosphate transport capability and the phosphorus storage capacity is higher in S. Nageli than in S. quadricauda. Synechococcus Nageli can also deplete phosphate to much lower levels than can S. quadricauda. These results, along with their morphological characteristics, were used to construct partial physiological profiles for each organism. The profiles indicate that this unicellular cyanophyte (cyanobacterium) is better suited for growth in phosphorus-limited oligotrophic niches than is this chlorophyte (green alga). (Refs. 44).
  • The PrrBA two-component regulatory system is a major global regulator in Rhodobacter sphaeroides 2.4.1. In this study we have compared the transcriptome and proteome profiles of the wild type (WT) and mutant PrrA2 cells grown anaerobically, in the dark, with DMSO as electron acceptor. Approximately 25% of the genes present in the genome are PrrA-regulated, at the transcriptional level, either directly or indirectly, by ≥ 2-fold relative to wild type. The genes affected are widespread throughout all COG functional categories, with previously unsuspected “metabolic” genes affected when in the PrrA mutant background. PrrA was found to act both as anmore » activator and a repressor of transcription, with more genes being repressed in the presence of PrrA (9:5 ratio). An analysis of the genes encoding the 1,536 peptides detected through our chromatographic study, which corresponds to 36% coverage of the genome, revealed that approximately 20% of the genes encoding these proteins were positively regulated, whereas approximately 32% were negatively regulated by PrrA, which is in excellent agreement with the percentages obtained for the whole genomic transcriptome profile. In addition, comparison of the transcriptome and proteome mean parameter values chosen between WT and PrrA2 showed good qualitative agreement, indicating that transcript regulation paralleled the corresponding protein abundance, although not one for one. The microarray analysis was validated by direct mRNA measurement of randomly selected, both positively and negatively regulated genes. lacZ transcriptional and kan translational fusions enabled us to map putative PrrA binding sites, as well as revealing potential gene targets for indirect regulation by PrrA.« less