Human colon cell migration and socialization induced by small RNA nucleotides

Buechler, Andrew K

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https://hdl.handle.net/2142/117548 Copy

Description

Title

Human colon cell migration and socialization induced by small RNA nucleotides

Author(s)

Buechler, Andrew K

Issue Date

2022-12-02

Director of Research (if dissertation) or Advisor (if thesis)

Martinis, Susan A

Doctoral Committee Chair(s)

Martinis, Susan A

Committee Member(s)

• Chen, Lin-Feng
• Hergenrother, Paul J
• Kalsotra, Auinash

Department of Study

Biochemistry

Discipline

Biochemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• RNA
• nucleic acids
• signaling nucleotides
• microbiota
• cell aggregation
• cell migration
• colon

Abstract

Aminoacyl-tRNA synthetases and tRNA molecules are well-known as essential housekeeping factors for protein translation. Additionally, they have adapted through evolution to carry out critical tasks in other cellular processes. These roles include crucial cell signaling functions and can involve translocation from the cytoplasm into other compartments and to the extracellular space. The human body carries trillions of bacterial cells on its external and internal surfaces, collectively called the microbiota. With the members of the microbiota roughly equaling the number of human cells in a body, it is not surprising that they have an enormous impact on human physiology, health, and disease. This relationship between the microbiota and their human hosts is modulated by simple and complex molecules, including metabolites, antibiotics, and macromolecular proteins. Previously, it was determined that leucyl-tRNA synthetase and tRNALeu are selectively secreted from nutritionally stressed Escherichia coli. Because E. coli resides in the human colon, both factors were incubated with human colon carcinoma cells. Remarkably, the tRNA molecules triggered colon cell migration and assembly into tightly packed cell aggregates. This “socialization” was reversible and dependent on nutritional deprivation. The structural characteristics of active RNA signals in socialization were investigated. Extracellular RNAs at least 15 nucleotides in length triggered socialization, regardless of sequence or secondary structural characteristics. However, chemically related molecules, such as DNA and 2’-O-methyl RNA, did not induce the same socialization phenotype. It was determined that any sufficiently long, 2’-hydroxyl-containing RNA would induce socialization as long as it was present in sufficient concentration, which varied based on its length. It was determined that these extracellular RNAs are processed into very short nucleotides before they trigger socialization. The active small nucleotides contain 2’- or 3’- terminal phosphates. One or more of these mono-, di-, and short oligonucleotides induce socialization, with guanosine-containing RNAs exhibiting the highest activity. Click chemistry fluorescence labeling of RNAs was employed to track the location of nucleotides over time. These experiments revealed that human colon cells internalize the RNA signaling factor. Upon entering cells, the RNA appears to be localized to the Golgi apparatus. With chemical disruption of the Golgi, the labelled nucleotide fragments migrate to the nucleus. Hundreds of genes were found to be differentially expressed during socialization via RNA-sequencing. Many of these genes are associated with innate immunity, cancer growth, and metastasis. In particular, these genes were enriched for the KEGG pathways that include p53 signaling, cell adhesion molecules, cellular senescence, gap junctions, and tight junctions. We hypothesize that tRNA molecules have been repurposed to influence the multi-faceted relationship between bacteria and their human hosts.

Graduation Semester

2022-12

Type of Resource

Thesis

Copyright and License Information

Copyright 2022 Andrew Buechler

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