Article

A role for lengsin, a recruited enzyme, in terminal differentiation in the vertebrate lens

National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Journal of Biological Chemistry (Impact Factor: 4.57). 04/2008; 283(10):6607-15. DOI: 10.1074/jbc.M709144200
Source: PubMed

ABSTRACT Lengsin is an eye lens-specific member of the glutamine synthetase (GS) superfamily. Lengsin has no GS activity, suggesting that it has a structural rather than catalytic role in lens. In situ hybridization and immunofluorescence showed that lengsin is expressed in terminally differentiating secondary lens fiber cells. Yeast two-hybrid (Y2H) and recombinant protein experiments showed that full-length lengsin can bind the 2B filament region of vimentin. In affinity chromatography, lengsin also bound the equivalent region of CP49 (BFSP2; phakinin), a related intermediate filament protein specific to the lens. Both the vimentin and CP49 2B fragments bound lengsin in surface plasmon resonance spectroscopy with fast association and slow dissociation kinetics. Lengsin expression correlates with a transition zone in maturing lens fiber cells in which cytoskeleton is reorganized. Lengsin and lens intermediate filament proteins co-localize at the plasma membrane in maturing fiber cells. This suggests that lengsin may act as a component of the cytoskeleton itself or as a chaperone for the reorganization of intermediate filament proteins during terminal differentiation in the lens.

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    • "), aA-crystallin (CRYAA) (Hawse et al. 2005), and aB-crystallin (Hawse et al. 2005); (F) actin-capping regulator encoding transcript tropomodulin 1 (TMOD1) (Nowak and Fowler 2012); (G) Lensgin (GLULD1/LGSN) (Wyatt et al. 2008); (H) cell-cycle regulator encoding transcript cyclin-dependent kinase 2 (CDK2) (Gao et al. 1999); (I) lens signaling encoding transcripts coiled-coil domain containing 80 (CCDC80/EQUARIN) (Song et al. 2012), EPH receptor type A2 (EPHA2) (Shi et al. 2012; Cheng et al. 2013), fibroblast growth factor receptor 2 (FGFR2) (Robinson 2006), and frizzled class receptor 3 (FZD3) (Dawes et al. 2013); and (J) lens DNA binding encoding transcripts paired box 6 (PAX6) (Cvekl and Piatigorsky 1996), heat shock transcription factor 4 (HSF4) (Fujimoto et al. 2004; Somasundaram and Bhat 2004), SRY (sex determining region Y)-box 2 (SOX2) (Kondoh et al. 2004), prospero homeobox 1 (PROX1) (Duncan et al. 2002), and GATA binding protein 3 (GATA3) (Maeda et al. 2009). "
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    ABSTRACT: The mature eye lens contains a surface layer of epithelial cells called the lens epithelium that require a functional mitochondrial population to maintain the homeostasis and transparency of the entire lens. The lens epithelium overlies a core of terminally differentiated fiber cells that must degrade their mitochondria to achieve lens transparency. These distinct mitochondrial populations make the lens a useful model system to identify those genes that regulate the balance between mitochondrial homeostasis and elimination. Here we used an RNA sequencing and bioinformatics approach to identify the transcript levels of all genes expressed by distinct regions of the lens epithelium and maturing fiber cells of the embryonic Gallus gallus (chicken) lens. Our analysis detected over 15,000 unique transcripts expressed by the embryonic chicken lens. Of these, over 3000 transcripts exhibited significant differences in expression between lens epithelial cells and fiber cells. Multiple transcripts coding for separate mitochondrial homeostatic and degradation mechanisms were identified to exhibit preferred patterns of expression in lens epithelial cells that require mitochondria relative to lens fiber cells that require mitochondrial elimination. These included differences in the expression levels of metabolic (DUT, PDK1, SNPH), autophagy (ATG3, ATG4B, BECN1, FYCO1, WIPI1), and mitophagy (BNIP3L/NIX, BNIP3, PARK2, p62/SQSTM1) transcripts between lens epithelial cells and lens fiber cells. These data provide a comprehensive window into all genes transcribed by the lens and those mitochondrial regulatory and degradation pathways that function to maintain mitochondrial populations in the lens epithelium and to eliminate mitochondria in maturing lens fiber cells.
    G3-Genes Genomes Genetics 06/2014; 4(8). DOI:10.1534/g3.114.012120 · 2.51 Impact Factor
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    • "7B, C). Furthermore, Lengsin is expressed in the subpopulation of early differentiating fiber cells which are not yet denucleated (Fig. 7D; (Harding et al., 2008; Wyatt et al., 2008)). In uhrf1 and dnmt1 mutants, Lengsin is expressed in the disorganized cells that make up the mutant lens periphery (Figs. "
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    ABSTRACT: DNA methylation is one of the key mechanisms underlying the epigenetic regulation of gene expression. During DNA replication, the methylation pattern of the parent strand is maintained on the replicated strand through the action of Dnmt1 (DNA Methyltransferase 1). In mammals, Dnmt1 is recruited to hemimethylated replication foci by Uhrf1 (Ubiquitin-like, Containing PHD and RING Finger Domains 1). Here we show that Uhrf1 is required for DNA methylation in vivo during zebrafish embryogenesis. Due in part to the early embryonic lethality of Dnmt1 and Uhrf1 knockout mice, roles for these proteins during lens development have yet to be reported. We show that zebrafish mutants in uhrf1 and dnmt1 have defects in lens development and maintenance. uhrf1 and dnmt1 are expressed in the lens epithelium, and in the absence of Uhrf1 or of catalytically active Dnmt1, lens epithelial cells have altered gene expression and reduced proliferation in both mutant backgrounds. This is correlated with a wave of apoptosis in the epithelial layer, which is followed by apoptosis and unraveling of secondary lens fibers. Despite these disruptions in the lens fiber region, lens fibers express appropriate differentiation markers. The results of lens transplant experiments demonstrate that Uhrf1 and Dnmt1 functions are required lens-autonomously, but perhaps not cell-autonomously, during lens development in zebrafish. These data provide the first evidence that Uhrf1 and Dnmt1 function is required for vertebrate lens development and maintenance.
    Developmental Biology 02/2011; 350(1):50-63. DOI:10.1016/j.ydbio.2010.11.009 · 3.64 Impact Factor
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    • "The change in nuclear shape may be secondary to dismantling of the nuclear lamina which, according to studies in bovine (Dahm et al., 1998) and chicken (Bassnett and Mataic, 1997) lenses, occurs before frank disintegration of the chromatin commences. Fiber cells are transcriptionally active until quite late in differentiation , as evidenced by their ability to transcribe novel mRNAs, such as those encoding lengsin (Wyatt et al., 2008) and hop (Vasiliev et al., 2007). Microinjection of expression plasmids into fiber cells located at various depths from the lens surface has confirmed that the majority of nucleated fiber cells are transcriptionally competent (Shestopalov and Bassnett, 1999). "
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    ABSTRACT: The programmed elimination of cytoplasmic organelles occurs during terminal differentiation of erythrocytes, keratinocytes and lens fiber cells. In each case, the process is relatively well understood phenomenologically, but the underlying molecular mechanisms have been surprisingly slow to emerge. This brief review considers the particular case of the lens where, in addition to their specialized physiological roles, organelles represent potential sources of light scattering. The article describes how the elimination of organelles from lens cells located on the visual axis contributes to the transparency of lens tissue. Classic anatomical studies of lens organelle degradation are discussed, along with more contemporary work utilizing confocal microscopy and other imaging modalities. Finally, recent data on the biochemistry of organelle degradation are reviewed. Several review articles on lens organelle degradation are available [Wride, M.A., 1996. Cellular and molecular features of lens differentiation: a review of recent advances. Differentiation 61, 77–93; Wride, M.A., 2000. Minireview: apoptosis as seen through a lens. Apoptosis 5, 203–209; Bassnett, S., 2002. Lens organelle degradation. Exp. Eye Res. 74, 1–6; Dahm, R., 2004. Dying to see. Sci. Am. 291, 82–89] and readers are directed to these for a comprehensive discussion of the earlier literature on this topic.
    Experimental Eye Research 02/2009; 88(2-88):133-139. DOI:10.1016/j.exer.2008.08.017 · 3.02 Impact Factor
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