Proteasome activity in a naïve mosquito cell line infected with Wolbachia pipientis wAlbB.
We used Wolbachia pipientis strain wAlbB from Aedes albopictus Aa23 cells to infect clonal Ae. albopictus TK-6 cells, which are resistant to 5-bromodeoxyuridine. Infected TK-6 cells were cultured in medium containing 5-bromodeoxyuridine to select against Aa23 cells that might have persisted in the inoculum. Infected TK-6 lines retained the Wolbachia infection for 5 mo, indicating that their metabolic processes support Wolbachia growth and multiplication. To investigate early events after Wolbachia infection, we labeled infected cells with (35)S[methionine/cysteine]. Patterns of labeled proteins on sodium dodecyl sulfate gels were similar in control and infected cells, with the exception of a 29-kDa protein. Tandem mass spectrometry revealed that the 29-kDa band included alpha and beta subunits of the 26S proteasome. Independent confirmation of the up-regulation of the proteasome was established by probing Western blots with a monoclonal antibody to the proteasome-associated co-factor, ubiquitin. Wolbachia's loss of metabolic pathways for the synthesis of most amino acids and retention of pathways for their uptake and metabolism suggest that proteasome activation provides a mechanism whereby controlled degradation of intracellular host proteins would increase availability of amino acids to support establishment and maintenance of the Wolbachia infection.
Fallon
Ann M
AM
Department of Entomology, University of Minnesota, St. Paul, MN 55108, USA. fallo002@umn.edu
Witthuhn
Bruce A
BA
D000330:Aedes / Q000187:drug effects / Q000378:metabolism* / Q000382:microbiology*; D000595:Amino Acid Sequence; D000818:Animals; D001973:Bromodeoxyuridine / Q000494:pharmacology; D002460:Cell Line; D008969:Molecular Sequence Data; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*; D054875:Ubiquitination / Q000502:physiology; D015854:Up-Regulation / Q000502:physiology; D020577:Wolbachia / Q000187:drug effects / Q000254:growth & development* / Q000378:metabolism*
D025801:Ubiquitin; D046988:Proteasome Endopeptidase Complex; D001973:Bromodeoxyuridine