Coordinate control of cell cycle regulatory genes in zebrafish development tested by cyclin D1 knockdown with morpholino phosphorodiamidates and hydroxyprolyl-phosphono peptide nucleic acids

Kevin T. Duffy, Mary Frances McAleer, William R. Davidson, Laszlo Kari, Csaba Kari, Chang Gong Liu, Steven A. Farber, Keith C. Cheng, Jason R. Mest, Eric Wickstrom, Adam P. Dicker, Ulrich Rodeck

Research output: Contribution to journalArticle

31 Citations (Scopus)

Abstract

During early zebrafish (Danio rerio) development zygotic transcription does not begin until the mid-blastula transition (MBT) ∼3 h after fertilization. MBT demarcates transition from synchronous short cell cycles of S and M phases exclusively to full cycles encompassing G1 and G2 phases. Transcriptional profiling and RT-PCR analyses during these phases enabled us to determine that this shift corresponds to decreased transcript levels of S/M phase cell cycle control genes (e.g. ccna2, ccnb1, ccnb2 and ccne) and increased transcript levels of ccnd1, encoding cyclin D1, and orthologs of p21 (p21-like) and retinoblastoma (Rb-like 1). To investigate the regulation of this process further, the translation of ccnd1 mRNA, a G1/S checkpoint control element, was impaired by microinjection of ccnd1-specific morpholino phosphorodiamidate (MO) 20mer or hydroxyprolyl-phosphono peptide nucleic acid (HypNA-pPNA) 16mer antisense oligonucleotides. The resulting downregulation of cyclin D1 protein resulted in microophthalmia and microcephaly, but not lethality. The phenotypes were not seen with 3-mismatch MO 20mers or 1-mismatch HypNA-pPNA 16mers, and were rescued by an exogenous ccnd1 mRNA construct with five mismatches. Collectively, these results indicate that transcription of key molecular determinants of asynchronous cell cycle control in zebrafish embryos commences at MBT and that the reduction of cyclin D1 expression compromises zebrafish eye and head development.

Original languageEnglish (US)
Pages (from-to)4914-4921
Number of pages8
JournalNucleic acids research
Volume33
Issue number15
DOIs
StatePublished - Oct 17 2005

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Peptide Nucleic Acids
cdc Genes
Morpholinos
Cyclin D1
Zebrafish
Regulator Genes
Blastula
Cell Cycle Checkpoints
S Phase
Cell Division
Microcephaly
Retinoblastoma
Antisense Oligonucleotides
G2 Phase
G1 Phase
Microinjections
Protein Biosynthesis
Fertilization
Cell Cycle
Down-Regulation

ASJC Scopus subject areas

  • Genetics

Cite this

Coordinate control of cell cycle regulatory genes in zebrafish development tested by cyclin D1 knockdown with morpholino phosphorodiamidates and hydroxyprolyl-phosphono peptide nucleic acids. / Duffy, Kevin T.; McAleer, Mary Frances; Davidson, William R.; Kari, Laszlo; Kari, Csaba; Liu, Chang Gong; Farber, Steven A.; Cheng, Keith C.; Mest, Jason R.; Wickstrom, Eric; Dicker, Adam P.; Rodeck, Ulrich.

In: Nucleic acids research, Vol. 33, No. 15, 17.10.2005, p. 4914-4921.

Research output: Contribution to journalArticle

Duffy, Kevin T. ; McAleer, Mary Frances ; Davidson, William R. ; Kari, Laszlo ; Kari, Csaba ; Liu, Chang Gong ; Farber, Steven A. ; Cheng, Keith C. ; Mest, Jason R. ; Wickstrom, Eric ; Dicker, Adam P. ; Rodeck, Ulrich. / Coordinate control of cell cycle regulatory genes in zebrafish development tested by cyclin D1 knockdown with morpholino phosphorodiamidates and hydroxyprolyl-phosphono peptide nucleic acids. In: Nucleic acids research. 2005 ; Vol. 33, No. 15. pp. 4914-4921.
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abstract = "During early zebrafish (Danio rerio) development zygotic transcription does not begin until the mid-blastula transition (MBT) ∼3 h after fertilization. MBT demarcates transition from synchronous short cell cycles of S and M phases exclusively to full cycles encompassing G1 and G2 phases. Transcriptional profiling and RT-PCR analyses during these phases enabled us to determine that this shift corresponds to decreased transcript levels of S/M phase cell cycle control genes (e.g. ccna2, ccnb1, ccnb2 and ccne) and increased transcript levels of ccnd1, encoding cyclin D1, and orthologs of p21 (p21-like) and retinoblastoma (Rb-like 1). To investigate the regulation of this process further, the translation of ccnd1 mRNA, a G1/S checkpoint control element, was impaired by microinjection of ccnd1-specific morpholino phosphorodiamidate (MO) 20mer or hydroxyprolyl-phosphono peptide nucleic acid (HypNA-pPNA) 16mer antisense oligonucleotides. The resulting downregulation of cyclin D1 protein resulted in microophthalmia and microcephaly, but not lethality. The phenotypes were not seen with 3-mismatch MO 20mers or 1-mismatch HypNA-pPNA 16mers, and were rescued by an exogenous ccnd1 mRNA construct with five mismatches. Collectively, these results indicate that transcription of key molecular determinants of asynchronous cell cycle control in zebrafish embryos commences at MBT and that the reduction of cyclin D1 expression compromises zebrafish eye and head development.",
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AU - McAleer, Mary Frances

AU - Davidson, William R.

AU - Kari, Laszlo

AU - Kari, Csaba

AU - Liu, Chang Gong

AU - Farber, Steven A.

AU - Cheng, Keith C.

AU - Mest, Jason R.

AU - Wickstrom, Eric

AU - Dicker, Adam P.

AU - Rodeck, Ulrich

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AB - During early zebrafish (Danio rerio) development zygotic transcription does not begin until the mid-blastula transition (MBT) ∼3 h after fertilization. MBT demarcates transition from synchronous short cell cycles of S and M phases exclusively to full cycles encompassing G1 and G2 phases. Transcriptional profiling and RT-PCR analyses during these phases enabled us to determine that this shift corresponds to decreased transcript levels of S/M phase cell cycle control genes (e.g. ccna2, ccnb1, ccnb2 and ccne) and increased transcript levels of ccnd1, encoding cyclin D1, and orthologs of p21 (p21-like) and retinoblastoma (Rb-like 1). To investigate the regulation of this process further, the translation of ccnd1 mRNA, a G1/S checkpoint control element, was impaired by microinjection of ccnd1-specific morpholino phosphorodiamidate (MO) 20mer or hydroxyprolyl-phosphono peptide nucleic acid (HypNA-pPNA) 16mer antisense oligonucleotides. The resulting downregulation of cyclin D1 protein resulted in microophthalmia and microcephaly, but not lethality. The phenotypes were not seen with 3-mismatch MO 20mers or 1-mismatch HypNA-pPNA 16mers, and were rescued by an exogenous ccnd1 mRNA construct with five mismatches. Collectively, these results indicate that transcription of key molecular determinants of asynchronous cell cycle control in zebrafish embryos commences at MBT and that the reduction of cyclin D1 expression compromises zebrafish eye and head development.

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