1. Biochemistry and Chemical Biology
Download icon

Long-lived metabolic enzymes in the Crystallin lens identified by pulse-labeling of mice and mass spectrometry

  1. Pan Liu
  2. Seby Louis Edassery
  3. Laith Ali
  4. Benjamin R Thomson
  5. Jeffrey N Savas  Is a corresponding author
  6. Jing Jin  Is a corresponding author
  1. Northwestern University, United States
Short Report
  • Cited 6
  • Views 1,032
  • Annotations
Cite this article as: eLife 2019;8:e50170 doi: 10.7554/eLife.50170

Abstract

The lenticular fiber cells are comprised of extremely long-lived proteins while still maintaining an active biochemical state. Dysregulation of these activities has been implicated in diseases such as age-related cataracts. However, the lenticular protein dynamics underlying health and disease is unclear. We sought to measure the global protein turnover rates in the eye using nitrogen-15 labeling of mice and mass spectrometry. We measured the 14N/15N-peptide ratios of 248 lens proteins, including Crystallin, Aquaporin, Collagen and enzymes that catalyze glycolysis and oxidation/reduction reactions. Direct comparison of lens cortex versus nucleus revealed little or no 15N-protein contents in most nuclear proteins, while there were a broad range of 14N/15N ratios in cortex proteins. Unexpectedly, like Crystallins, many enzymes with relatively high abundance in nucleus were also exceedingly long-lived. The slow replacement of these enzymes in spite of young age of mice suggests their potential roles in age-related metabolic changes in the lens.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data file (Supplementary data set) have been provided.

The following data sets were generated

Article and author information

Author details

  1. Pan Liu

    Feinberg Cardiovascular and Renal Research Institute, Feinberg School of Medicine, Northwestern University, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3066-652X

  2. Seby Louis Edassery

    Department of Neurology, Feinberg School of Medicine, Northwestern University, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Laith Ali

    Department of Neurology, Feinberg School of Medicine, Northwestern University, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Benjamin R Thomson

    Feinberg Cardiovascular and Renal Research Institute, Feinberg School of Medicine, Northwestern University, Chicago, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6565-5866

  5. Jeffrey N Savas

    Department of Neurology, Feinberg School of Medicine, Northwestern University, Chicago, United States
    For correspondence
    jeffrey.savas@northwestern.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8173-5580

  6. Jing Jin

    Feinberg Cardiovascular and Renal Research Institute, Feinberg School of Medicine, Northwestern University, Chicago, United States
    For correspondence
    jing.jin@northwestern.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7023-5305

Funding

National Institutes of Health (R01AG061787)

  • Jeffrey N Savas

National Institutes of Health (R21AI131087)

  • Jing Jin

National Institutes of Health (R01EY025799)

  • Jing Jin

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: All animal procedures were approved by Institutional Animal Care and Use Committee of the Northwestern University (approved protocol number IS00000429 and IS00000862).

Reviewing Editor

  1. Jeremy Nathans, Johns Hopkins University School of Medicine, United States

Publication history

  1. Received: July 12, 2019
  2. Accepted: December 4, 2019
  3. Accepted Manuscript published: December 10, 2019 (version 1)
  4. Version of Record published: December 16, 2019 (version 2)

Copyright

© 2019, Liu et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 1,032
    Page views
  • 137
    Downloads
  • 6
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Categories and tags

Research organism

Further reading

    1. Biochemistry and Chemical Biology

    Myopalladin knockout mice develop cardiac dilation and show a maladaptive response to mechanical pressure overload

    Maria Carmela Filomena et al.
    Research Article

    Myopalladin (MYPN) is a striated muscle-specific immunoglobulin domain-containing protein located in the sarcomeric Z-line and I-band. MYPN gene mutations are causative for dilated (DCM), hypertrophic and restrictive cardiomyopathy. In a yeast two-hybrid screening, MYPN was found to bind to titin in the Z-line, which was confirmed by microscale thermophoresis. Cardiac analyses of MYPN knockout (MKO) mice showed the development of mild cardiac dilation and systolic dysfunction, associated with decreased myofibrillar isometric tension generation and increased resting tension at longer sarcomere lengths. MKO mice exhibited a normal hypertrophic response to transaortic constriction (TAC), but rapidly developed severe cardiac dilation and systolic dysfunction, associated with fibrosis, increased fetal gene expression, higher intercalated disc fold amplitude, decreased calsequestrin-2 protein levels, and increased desmoplakin and SORBS2 protein levels. Cardiomyocyte analyses showed delayed Ca2+ release and reuptake in unstressed MKO mice as well as reduced Ca2+ spark amplitude post-TAC, suggesting that altered Ca2+ handling may contribute to the development of DCM in MKO mice.

    1. Biochemistry and Chemical Biology

    Witnessing the structural evolution of an RNA enzyme

    Xavier Portillo et al.
    Research Article Updated

    An RNA polymerase ribozyme that has been the subject of extensive directed evolution efforts has attained the ability to synthesize complex functional RNAs, including a full-length copy of its own evolutionary ancestor. During the course of evolution, the catalytic core of the ribozyme has undergone a major structural rearrangement, resulting in a novel tertiary structural element that lies in close proximity to the active site. Through a combination of site-directed mutagenesis, structural probing, and deep sequencing analysis, the trajectory of evolution was seen to involve the progressive stabilization of the new structure, which provides the basis for improved catalytic activity of the ribozyme. Multiple paths to the new structure were explored by the evolving population, converging upon a common solution. Tertiary structural remodeling of RNA is known to occur in nature, as evidenced by the phylogenetic analysis of extant organisms, but this type of structural innovation had not previously been observed in an experimental setting. Despite prior speculation that the catalytic core of the ribozyme had become trapped in a narrow local fitness optimum, the evolving population has broken through to a new fitness locale, raising the possibility that further improvement of polymerase activity may be achievable.

    1. Biochemistry and Chemical Biology

    A universal pocket in fatty acyl-AMP ligases ensures redirection of fatty acid pool away from coenzyme A-based activation

    Gajanan S Patil et al.
    Research Article Updated

    Fatty acyl-AMP ligases (FAALs) channelize fatty acids towards biosynthesis of virulent lipids in mycobacteria and other pharmaceutically or ecologically important polyketides and lipopeptides in other microbes. They do so by bypassing the ubiquitous coenzyme A-dependent activation and rely on the acyl carrier protein-tethered 4′-phosphopantetheine (holo-ACP). The molecular basis of how FAALs strictly reject chemically identical and abundant acceptors like coenzyme A (CoA) and accept holo-ACP unlike other members of the ANL superfamily remains elusive. We show that FAALs have plugged the promiscuous canonical CoA-binding pockets and utilize highly selective alternative binding sites. These alternative pockets can distinguish adenosine 3′,5′-bisphosphate-containing CoA from holo-ACP and thus FAALs can distinguish between CoA and holo-ACP. These exclusive features helped identify the omnipresence of FAAL-like proteins and their emergence in plants, fungi, and animals with unconventional domain organizations. The universal distribution of FAALs suggests that they are parallelly evolved with FACLs for ensuring a CoA-independent activation and redirection of fatty acids towards lipidic metabolites.