Comparing protein stabilities during zebrafish embryogenesis.
The stabilities of many key proteins are regulated, e.g. via ubiquitination and proteasomal degradation, with important biological consequences. We present a convenient method that allows the analysis and comparison of protein stabilities during embryogenesis using early zebrafish development as a model system. Basically, this method involves ectopic overexpression of epitope-tagged proteins via mRNA injections in one-to-four-cell stage embryos and subsequent protein detection after various time points. Indeed, the protein stability of the ubiquitin ligase RLIM, which is able to autoubiquitinate and target itself for proteasomal degradation, was much shorter when compared to a protein consisting of a Myc epitope-tag and a nuclear localization domain. Thus, this method may be used more widely for the study of developmental protein stability.
Becker
Thomas
T
Zentrum für Molekulare Neurobiologie (ZMNH), Universität Hamburg, Martinistr. 85, 20251 Hamburg, Germany.
Bossenz
Michael
M
Tursun
Baris
B
Schlüter
Anne
A
Peters
Marvin A
MA
Becker
Catherina G
CG
Ostendorff
Heather P
HP
Bach
Ingolf
I
D000818:Animals; D003001:Cloning, Molecular; D004625:Embryo, Nonmammalian / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D008845:Microinjections; D011489:Protein Denaturation; D020871:RNA Stability / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism*; D015027:Zebrafish / Q000196:embryology / Q000378:metabolism*
D012333:RNA, Messenger; D012097:Repressor Proteins