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Related Concept Videos

Ribosome Profiling02:24

Ribosome Profiling

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Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique...
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Ribosomes01:27

Ribosomes

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Ribosomes translate genetic information encoded by messenger RNA (mRNA) into proteins. Both prokaryotic and eukaryotic cells have ribosomes. Cells that synthesize large quantities of protein—such as secretory cells in the human pancreas—can contain millions of ribosomes.
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Restarting Stalled Replication Forks02:37

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DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart,...
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Initiation of Translation02:33

Initiation of Translation

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Initiating translation is complex because it involves multiple molecules. Initiator tRNA, ribosomal subunits, and eukaryotic initiation factors (eIFs) are all required to assemble on the initiation codon of mRNA. This process consists of several steps that are mediated by different eIFs.
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Ribosomal RNA Synthesis02:53

Ribosomal RNA Synthesis

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Ribosome synthesis is a highly complex and coordinated process involving more than 200 assembly factors. The synthesis and processing of ribosomal components occurs not only in the nucleolus but also in the nucleoplasm and the cytoplasm of eukaryotic cells.
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Protein Complex Assembly02:41

Protein Complex Assembly

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Proteins can form homomeric complexes with another unit of the same protein or heteromeric complexes with different types.  Most protein complexes self-assemble spontaneously via ordered pathways, while some proteins need assembly factors that guide their proper assembly. Despite the crowded intracellular environment, proteins usually interact with their correct partners and form functional complexes.
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De novo Identification of Actively Translated Open Reading Frames with Ribosome Profiling Data
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De novo Identification of Actively Translated Open Reading Frames with Ribosome Profiling Data

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Identifying Small Proteins by Ribosome Profiling with Stalled Initiation Complexes.

Jeremy Weaver1, Fuad Mohammad2, Allen R Buskirk3

  • 1Division of Molecular and Cellular Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, Maryland, USA.

Mbio
|March 7, 2019
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Summary

Researchers discovered 38 new small proteins in E. coli using ribosome profiling. This method identifies small proteins missed by traditional methods, revealing their roles in gene regulation.

Keywords:
Ribo-seqalternate ORFsantisensegenome annotationleader peptidesmall protein

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Area of Science:

  • Microbiology
  • Molecular Biology
  • Genomics

Background:

  • Small proteins (<50 amino acids) regulate larger proteins in bacteria and eukaryotes.
  • Conventional methods often miss these small proteins due to their small size, excluding small open reading frames (smORFs).
  • Previous work identified some small proteins in E. coli using theoretical bioinformatic approaches.

Purpose of the Study:

  • To develop and apply an empirical method for discovering novel small proteins.
  • To identify new small proteins in Escherichia coli.
  • To characterize the genomic context and potential functions of newly identified small proteins.

Main Methods:

  • Utilized ribosome profiling with antibiotics to trap initiating 70S ribosomes at start codons.
  • Identified novel initiation sites in intergenic regions of E. coli.
  • Experimentally tagged and confirmed protein synthesis for 41 small open reading frames (smORFs).

Main Results:

  • Identified numerous novel initiation sites, leading to the discovery of 38 new small proteins.
  • Found that these small protein genes are often intergenic, antisense, in operons, or overlapping other open reading frames (ORFs).
  • Observed that some small proteins impact the translation of downstream genes.

Conclusions:

  • Ribosome profiling with stalled initiation complexes is an effective empirical method for discovering small proteins.
  • The identified small proteins, despite their varied genomic locations, play regulatory roles.
  • This approach significantly expands the known proteome by revealing previously hidden small proteins.