Dataset Information

Birc7: A Late Fiber Gene of the Crystalline Lens.

ABSTRACT:

Purpose

A distinct subset of genes, so-called "late fiber genes," is expressed in cells bordering the central, organelle-free zone (OFZ) of the lens. The purpose of this study was to identify additional members of this group.

Methods

Fiber cells were harvested from various layers of the lens by laser micro-dissection and subjected to microarray, in situ hybridization, and Western blot analysis.

Results

Expression of Livin, a member of the inhibitor of apoptosis protein (IAP) family encoded by Birc7, was strongly upregulated in deep cortical fiber cells. The depth-dependent distribution of Livin mRNA was confirmed by quantitative PCR and in situ hybridization. The onset of Livin expression coincided with loss of organelles from primary fiber cells. Livin expression peaked at 1 month but was sustained even in aged lenses. Antibodies raised against mouse Livin labeled multiple bands on immunoblots, reflecting progressive proteolysis of the parent molecule during differentiation. Mice harboring a floxed Birc7 allele were generated and used to conditionally delete Birc7 in lens. Lenses from knockout mice grew normally and retained their transparency, suggesting that Livin does not have an indispensable role in fiber cell differentiation.

Conclusions

Birc7 is a late fiber gene of the mouse lens. In tumor cells, Livin acts as an antiapoptotic protein, but its function in the lens is enigmatic. Livin is a RING domain protein with putative E3 ubiquitin ligase activity. Its expression in cells bordering the OFZ is consistent with a role in organelle degradation, a process in which the ubiquitin proteasome pathway has been implicated previously.

SUBMITTER: De Maria A

PROVIDER: S-EPMC4520384 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Publications

Birc7: A Late Fiber Gene of the Crystalline Lens.

De Maria Alicia A Bassnett Steven S

Investigative ophthalmology & visual science 20150701 8

<h4>Purpose</h4>A distinct subset of genes, so-called "late fiber genes," is expressed in cells bordering the central, organelle-free zone (OFZ) of the lens. The purpose of this study was to identify additional members of this group.<h4>Methods</h4>Fiber cells were harvested from various layers of the lens by laser micro-dissection and subjected to microarray, in situ hybridization, and Western blot analysis.<h4>Results</h4>Expression of Livin, a member of the inhibitor of apoptosis protein (IAP ...[more]

PMID: 26218911

Similar Datasets

Project description:To identify transcripts expressed late in lens fiber cell maturation that might regulate fiber cell fusion, organelle degradation, or other events associated with the maturation of lens fiber cells.cDNA libraries were prepared from microdissected regions of chicken embryo lenses using a PCR-based method. Subtractive hybridization was used to identify transcripts expressed exclusively in fiber cells that had detached from the lens capsule. Database searches and PCR amplification with degenerate primers were used to identify human, mouse, rat, rabbit, and bovine orthologs of one such sequence and to confirm its expression in the lenses of these animals. The ability of in vitro-transcribed and translated protein to bind DNA was assessed by mobility shift assays. The locus encoding this transcript and an area about 6 kb upstream of the translation start site were sequenced. The microscopic morphology of lenses from mice in which the locus encoding this protein had been disrupted by the insertion of a nuclear-targeted bacterial lacZ sequence were analyzed. Gene expression was analyzed by PCR, in situ hybridization, and by staining for beta-galactosidase activity in lenses expressing lacZ in place of the coding sequence. Knockout lenses expressing green fluorescent protein in a mosaic pattern were sectioned in the equatorial plane and viewed with a confocal microscope to assess the presence of cell-cell fusions during fiber cell maturation.Subtractive hybridization identified transcripts encoding Hop, a short, atypical homeodomain-containing protein that had previously been shown to be an important regulator of gene expression in the heart and lung. Chicken Hop did not bind to known homeodomain-binding sequences in DNA. In chicken embryos, Hop transcripts were first detected at E6. At all stages analyzed, Hop mRNA was only detected in cells that had detached from the lens capsule. Mice in which the Hop coding sequence was replaced with nuclear-targeted beta-galactosidase showed that Hop was expressed in the mouse lens in a similar pattern to the chicken lens. Characterization of lenses from mice lacking Hop revealed no morphological phenotype and no apparent defects in the degradation of nuclei or fiber cell fusion during fiber cell maturation.The expression pattern of Hop provides the first evidence that new transcription is initiated in lens fiber cells after they detach from the capsule. Hop may be the first of a class of genes with this pattern of expression. Although lens abnormalities have yet to be identified in mice lacking Hop, the genomic sequences that regulate Hop expression in the lens may be useful for expressing exogenous transcripts selectively in fiber cells just before they fuse with their neighbors and degrade their organelles.

Dataset Information

Birc7: A Late Fiber Gene of the Crystalline Lens.

Purpose

Methods

Results

Conclusions

Publications

Birc7: A Late Fiber Gene of the Crystalline Lens.

Similar Datasets

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