A ribosome-free extract from cultured cells improves recovery of polysomes from the mosquito fat body: Analysis of vitellogenin and ribosomal protein rpL34 gene expression

Ling Ling Niu, Ann M. Fallon

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

We have examined the association of ribosomal protein rpL34 mRNA with polysomes in Aedes albopictus C7-10 cells in culture using a simple, two-step sucrose gradient. In growing cells, 40-50% of the ribosomes were engaged on polysomes. This proportion could be increased to 80% when metabolism was stimulated by refeeding the cells with fresh medium. Conversely, ribosomes shifted off polysomes when cells were starved with phosphate-buffered saline or cell lysates were treated with puromycin. When similar approaches were used with fat body from blood-fed female Aedes aegypti mosquitoes, we were unable to obtain the polysome fraction that contained vitellogenin mRNA, which is abundantly translated after a blood meal. Addition of post-mitochondrial supernatant from fat body to polysomes from cultured cells shifted the polysome profile towards smaller polysomes and monosomes, in a dose-dependent fashion. Disruption of fat body tissue in a post-ribosomal supernatant from refed cells improved the recovery of polysomes, demonstrating both the engagement of vitellogenin mRNA on polysomes and the mobilization of rpL34 from messenger-ribonuceloprotein particles onto polysomes in blood-fed mosquitoes. These observations suggested that ribonucleases remain active when polysomes are prepared from mosquito fat body, and that cell culture supernatant contains a ribonuclease inhibitor.

Original languageEnglish (US)
Pages (from-to)835-843
Number of pages9
JournalJournal of Insect Physiology
Volume48
Issue number9
DOIs
StatePublished - Sep 1 2002

Bibliographical note

Funding Information:
This research was supported by NIH Grant No. AI 20385 and by the University of Minnesota Agricultural Experiment Station, St. Paul, MN.

Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.

Keywords

  • Fat body
  • Mosquito
  • Polysomes
  • Translational control
  • Vitellogenin

Fingerprint Dive into the research topics of 'A ribosome-free extract from cultured cells improves recovery of polysomes from the mosquito fat body: Analysis of vitellogenin and ribosomal protein rpL34 gene expression'. Together they form a unique fingerprint.

Cite this