PUBLICATION

Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas

Authors
Won, M., Ro, H., Dawid, I.B.
ID
ZDB-PUB-150924-11
Date
2015
Source
Proceedings of the National Academy of Sciences of the United States of America   112(40): 12426-31 (Journal)
Registered Authors
Dawid, Igor B., Ro, Hyunju, Won, Minho
Keywords
Notch, Numb, TALEN, morpholino, pancreas
MeSH Terms
  • Reverse Transcriptase Polymerase Chain Reaction
  • Gene Knockdown Techniques
  • Zebrafish Proteins/genetics*
  • Zebrafish Proteins/metabolism
  • Pancreas, Exocrine/cytology
  • Pancreas, Exocrine/embryology
  • Pancreas, Exocrine/metabolism*
  • Gene Expression Regulation, Developmental
  • Humans
  • Animals
  • Alternative Splicing
  • Ubiquitin-Protein Ligases/genetics*
  • Ubiquitin-Protein Ligases/metabolism
  • Immunoblotting
  • Microscopy, Confocal
  • Base Sequence
  • Amino Acid Sequence
  • Cell Differentiation/genetics*
  • HEK293 Cells
  • Morpholinos/genetics
  • In Situ Hybridization
  • Mutation
  • Zebrafish/embryology
  • Zebrafish/genetics
  • Zebrafish/metabolism
  • Carrier Proteins/genetics*
  • Carrier Proteins/metabolism
  • Pancreas/cytology
  • Pancreas/embryology
  • Pancreas/metabolism*
(all 30)
PubMed
26392552 Full text @ Proc. Natl. Acad. Sci. USA
Abstract
The gene encoding the E3 ubiquitin ligase Ligand of Numb protein-X (Lnx)2a is expressed in the ventral-anterior pancreatic bud of zebrafish embryos in addition to its expression in the brain. Knockdown of Lnx2a by using an exon 2/intron 2 splice morpholino resulted in specific inhibition of the differentiation of ventral bud derived exocrine cell types, with little effect on endocrine cell types. A frame shifting null mutation in lnx2a did not mimic this phenotype, but a mutation that removed the exon 2 splice donor site did. We found that Lnx2b functions in a redundant manner with its paralog Lnx2a. Inhibition of lnx2a exon 2/3 splicing causes exon 2 skipping and leads to the production of an N-truncated protein that acts as an interfering molecule. Thus, the phenotype characterized by inhibition of exocrine cell differentiation requires inactivation of both Lnx2a and Lnx2b. Human LNX1 is known to destabilize Numb, and we show that inhibition of Numb expression rescues the Lnx2a/b-deficient phenotype. Further, Lnx2a/b inhibition leads to a reduction in the number of Notch active cells in the pancreas. We suggest that Lnx2a/b function to fine tune the regulation of Notch through Numb in the differentiation of cell types in the early zebrafish pancreas. Further, the complex relationships among genotype, phenotype, and morpholino effect in this case may be instructive in the ongoing consideration of morpholino use.
Genes / Markers
Figures
Figure Gallery (13 images) / 2
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Expression
Gene Antibody Fish Conditions Stage Qualifier Anatomy Assay Figure
EGFPjh1TgcontrolHigh-pecIFL
EGFPjh1TgcontrolLong-pecIFL
EGFPjh1TgcontrolLong-pecIFL
EGFPjh1Tg + MO1-lnx2acontrolLong-pecIFL
EGFPjh1Tg + MO1-lnx2acontrolHigh-pecIFL
foxa3ABcontrolPrim-25ISH
foxa3ABcontrolLong-pecISH
foxa3ABcontrolPrim-25ISH
foxa3ABcontrolPrim-25ISH
foxa3AB + MO1-lnx2acontrolPrim-25ISH
1 - 10 of 148
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Phenotype
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
jh1TgTransgenic Insertion
    s939TgTransgenic Insertion
      s940TgTransgenic Insertion
        y447
          		Small Deletion
          y448
            		Small Deletion
            1 - 5 of 5
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            Human Disease / Model
            No data available
            Sequence Targeting Reagents
            Target Reagent Reagent Type
            lnx2aMO1-lnx2aMRPHLNO
            lnx2aTALEN1-lnx2aTALEN
            lnx2bMO4-lnx2bMRPHLNO
            numbMO4-numbMRPHLNO
            1 - 4 of 4
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            Fish
            Fish
            AB
            AB + MO1-lnx2a
            AB + MO1-lnx2a + MO4-numb
            AB + MO4-lnx2b
            AB + MO4-numb
            jh1Tg
            jh1Tg + MO1-lnx2a
            lnx2ay447/y447
            lnx2ay447/y447
            lnx2ay447/y447
            1 - 10 of 17
            Show
            Antibodies
            Orthology
            No data available
            Engineered Foreign Genes
            Marker Marker Type Name
            EGFPEFGEGFP
            mCherryEFGmCherry
            VenusEFGVenus
            1 - 3 of 3
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            Mapping
            No data available
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            Citation
            Won, M., Ro, H., Dawid, I.B. (2015) Lnx2 ubiquitin ligase is essential for exocrine cell differentiation in the early zebrafish pancreas. Proceedings of the National Academy of Sciences of the United States of America. 112(40):12426-31.