Gitlin Jonathan D.

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Gitlin
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Jonathan D.
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Author: Thomason, Rebecca T. ×

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  • Article
    Microvillar and ciliary defects in zebrafish lacking an actin-binding bioactive peptide amidating enzyme
    (Nature Publishing Group, 2018-03-14) Kumar, Dhivya ; Thomason, Rebecca T. ; Yankova, Maya ; Gitlin, Jonathan D. ; Mains, Richard E. ; Eipper, Betty A. ; King, Stephen M.
    The assembly of membranous extensions such as microvilli and cilia in polarized cells is a tightly regulated, yet poorly understood, process. Peptidylglycine α-amidating monooxygenase (PAM), a membrane enzyme essential for the synthesis of amidated bioactive peptides, was recently identified in motile and non-motile (primary) cilia and has an essential role in ciliogenesis in Chlamydomonas, Schmidtea and mouse. In mammalian cells, changes in PAM levels alter secretion and organization of the actin cytoskeleton. Here we show that lack of Pam in zebrafish recapitulates the lethal edematous phenotype observed in Pam−/− mice and reveals additional defects. The pam−/− zebrafish embryos display an initial striking loss of microvilli and subsequently impaired ciliogenesis in the pronephros. In multiciliated mouse tracheal epithelial cells, vesicular PAM staining colocalizes with apical actin, below the microvilli. In PAM-deficient Chlamydomonas, the actin cytoskeleton is dramatically reorganized, and expression of an actin paralogue is upregulated. Biochemical assays reveal that the cytosolic PAM C-terminal domain interacts directly with filamentous actin but does not alter the rate of actin polymerization or disassembly. Our results point to a critical role for PAM in organizing the actin cytoskeleton during development, which could in turn impact both microvillus formation and ciliogenesis.
  • Article
    Characterization of trace metal content in the developing zebrafish embryo
    (Public Library of Science, 2017-06-15) Thomason, Rebecca T. ; Pettiglio, Michael A. ; Herrera, Carolina ; Kao, Clara ; Gitlin, Jonathan D. ; Bartnikas, Thomas B.
    Trace metals are essential for health but toxic when present in excess. The maintenance of trace metals at physiologic levels reflects both import and export by cells and absorption and excretion by organs. The mechanism by which this maintenance is achieved in vertebrate organisms is incompletely understood. To explore this, we chose zebrafish as our model organism, as they are amenable to both pharmacologic and genetic manipulation and comprise an ideal system for genetic screens and toxicological studies. To characterize trace metal content in developing zebrafish, we measured levels of three trace elements, copper, zinc, and manganese, from the oocyte stage to 30 days post-fertilization using inductively coupled plasma mass spectrometry. Our results indicate that metal levels are stable until zebrafish can acquire metals from the environment and imply that the early embryo relies on maternal contribution of metals to the oocyte. We also measured metal levels in bodies and yolks of embryos reared in presence and absence of the copper chelator neocuproine. All three metals exhibited different relative abundances between yolks and bodies of embryos. While neocuproine treatment led to an expected phenotype of copper deficiency, total copper levels were unaffected, indicating that measurement of total metal levels does not equate with measurement of biologically active metal levels. Overall, our data not only can be used in the design and execution of genetic, physiologic, and toxicologic studies but also has implications for the understanding of vertebrate metal homeostasis.