Biology subjects

Lewis, M. R.

Publications and source records attributed to Lewis, M. R..

4 recordsLinked to original sources

bioRxiv2025

Rare codon translation regulates growth factor-dependent cell proliferation

In animal cells, growth factor signaling promotes both cell cycle progression and cell growth, but the connection between these two processes is not well understood. Here, we investigated whether cell cycling and cell growth are coupled through protein translation. Using ribosome profiling and mRNA sequencing we examined changes in translational activities in human Retinal Pigment Epithelial (RPE-1) cells as they entered the cell cycle in response to serum growth factor stimulation. We found that, in addition to mRNAs encoding factors in ribosome biogenesis and translational initiation, mRNAs encoding many DNA replication factors were translationally upregulated by serum. We also noted increases of snoRNAs and tRNAs, which facilitate translation. By analyzing the distribution of 21nt mRNA ribosome footprints, produced by stalled ribosomes that lack amino-acyl-tRNAs, we found that growth factor withdrawal promoted ribosome stalling preferentially at specific rare codons. DNA replication factor genes that were translationally upregulated by serum and essential for cell cycle progression were enriched in many of these rare codons, and the cognate tRNAs that read these codons were induced by growth signaling. The serum-dependent induction of translation was more mTOR-dependent than MEK-dependent. Our results support a novel regulatory mechanism wherein growth factor signaling promotes cell proliferation by inducing tRNAs that decode rare codons, which in turn promote translational elongation of mRNAs encoding DNA replication factors to accelerate G1/S progression. Significance StatementWe present a mechanistic explanation for the longstanding question of how cell cycle progression is coupled to cell growth. Our findings emphasize the role of translation machinery and reveal a "growth checkpoint" that ensures that DNA replication gene translation occurs only when growth signaling has activated the protein translation machinery. Our work complements the textbook model of the Growth Factor-Cyclin D-Rb-E2F transcriptional mechanism for cell cycle entry and is a fundamental advance for the fields of cell proliferation and growth factor signaling.

cell biology↗

bioRxiv2024

Mating and ecdysone signaling modify growth, metabolism, and digestive efficiency in the female Drosophila gut

Adaptive changes in organ size and physiology occur in most adult animals, but how these changes are regulated is not well understood. Previous research found that mating in Drosophila females drives not only increases in gut size and stem cell proliferation but also alters feeding behavior, intestinal gene expression, and whole-body lipid storage, suggesting altered gut metabolism. Here, we show that mating dramatically alters female gut metabolism and digestive function. In addition to promoting a preference for a high-protein diet, mating also altered levels of TCA cycle intermediates and fatty acids in the gut, increased total gut lipids and protein, reduced relative carbohydrate levels, and enhanced the efficiency of protein digestion relative to carbohydrate digestion. The expression of genes that mediate each of these metabolic processes was similarly altered. In addition, we noted the mating-dependent downregulation of oxidative stress response and autophagy genes. Mating-dependent increases in ecdysone signaling played an important role in re-programming many, but not all, of these changes in the female gut. This study contributes to our understanding of how steroid signaling alters gut physiology to adapt to the demands of reproduction.

physiology↗

bioRxiv2023

In vitro generation of megakaryocytes from engineered mouse embryonic stem cells

The in vitro differentiation of pluripotent stem cells into desired lineages enables mechanistic studies of cell transitions into more mature states that can provide insights into the design principles governing cell fate control. We are interested in reprogramming pluripotent stem cells with synthetic gene circuits to drive mouse embryonic stem cells (mESCs) down the hematopoietic lineage for the production of megakaryocytes, the progenitor cells for platelets. Here, we describe the methodology for growing and differentiating mESCs, in addition to inserting a transgene to observe its expression throughout differentiation. This entails four key methods: (1) growing and preparing mouse embryonic fibroblasts for supporting mESC growth and expansion, (2) growing and preparing OP9 feeder cells to support the differentiation of mESCs, (3) the differentiation of mESCs into megakaryocytes, and (4) utilizing an integrase mediated docking site to insert transgenes for their stable integration and expression throughout differentiation. Altogether, this approach demonstrates a streamline differentiation protocol that emphasizes the reprogramming potential of mESCs that can be used for future mechanistic and therapeutic studies of controlling cell fate outcomes.

synthetic biology↗

bioRxiv2021

Mass recalibration for desorption electrospray ionization mass spectrometry imaging using endogenous reference ions.

1BackgroundMass spectrometry imaging (MSI) data often consist of tens of thousands of mass spectra collected from a sample surface. During the time necessary to perform a single acquisition, it is likely that uncontrollable factors alter the validity of the initial mass calibration of the instrument, resulting in mass errors of magnitude significantly larger than their expected values. This phenomenon has a two-fold detrimental effect: a) it reduces the ability to interpret the results based on the observed signals, b) it can affect the quality of the observed signal spatial distributions. ResultsWe present a post-acquisition computational method capable of reducing the observed mass drift by up to 60 ppm in biological samples, exploiting the presence of typical molecules with a known mass-to-charge ratio. The procedure, tested on time-of-flight (TOF) and Orbitrap mass spectrometry analyzers interfaced to a desorption electrospray ionization (DESI) source, improves the molecular annotation quality and the spatial distributions of the detected ions. ConclusionThe presented method represents a robust and accurate tool for performing post-acquisition mass recalibration of DESI-MSI datasets and can help to increase the reliability of the molecular assignment and the data quality.

bioinformatics↗