X-ray crystallographic studies of RoAb13 bound to PIYDIN, a part of the N-terminal domain of C-C chemokine receptor 5.

C-C chemokine receptor 5 (CCR5) is a major co-receptor molecule used by HIV-1 to enter cells. This led to the hypothesis that stimulating an antibody response would block HIV with minimal toxicity. Here, X-ray crystallographic studies of the anti-CCR5 antibody RoAb13 together with two peptides were undertaken: one peptide is a 31-residue peptide containing the PIYDIN sequence and the other is the PIDYIN peptide alone, where PIYDIN is part of the N-terminal region of CCR5 previously shown to be important for HIV entry.

Hydrophobic Interface-Assisted Protein Crystallization: Theory and Experiment.

Macromolecular crystallization is crucial to a large number of scientific fields, including structural biology; drug design, formulation, and delivery; manufacture of biomaterials; and preparation of foodstuffs. The purpose of this study is to facilitate control of crystallization, by investigating hydrophobic interface-assisted protein crystallization both theoretically and experimentally. The application of hydrophobic liquids as nucleation promoters or suppressors has rarely been investigated, and provides an underused avenue to explore in protein crystallization.

Stat3 defines three populations of Müller glia and is required for initiating maximal müller glia proliferation in the regenerating zebrafish retina.

We analyzed the role of Stat3, Ascl1a, and Lin28a in Müller glia reentry into the cell cycle following damage to the zebrafish retina. Immunohistochemical analysis was employed to determine the temporal and spatial expression of Stat3 and Ascl1a proteins following rod and cone photoreceptor cell apoptosis. Stat3 expression was observed in all Müller glia, whereas Ascl1a expression was restricted to only the mitotic Müller glia.

Pax6a and Pax6b are required at different points in neuronal progenitor cell proliferation during zebrafish photoreceptor regeneration.

The light-damaged zebrafish retina results in the death of photoreceptor cells and the subsequent regeneration of the missing rod and cone cells. Photoreceptor regeneration initiates with asymmetric Müller glial cell division to produce neuronal progenitor cells, which amplify, migrate to the outer nuclear layer (ONL), and differentiate into both classes of photoreceptor cells. In this study, we examined the role of the Pax6 protein in regeneration.

CNTF induces photoreceptor neuroprotection and Müller glial cell proliferation through two different signaling pathways in the adult zebrafish retina.

Ciliary neurotrophic factor (CNTF) acts in several processes in the vertebrate retina, including neuroprotection of photoreceptors in the stressed adult retina and regulation of neuronal progenitor cell proliferation during retinal development. However, the signaling pathway it utilizes (Jak/Stat, MAPK, or Akt) in these processes is ambiguous. Because dark-adapted albino zebrafish exhibit light-induced rod and cone cell death and subsequently regenerate the lost photoreceptor cells, zebrafish should be a useful model to study the role of CNTF in both neuroprotection and neuronal progenitor cell proliferation.

Characterization of Müller glia and neuronal progenitors during adult zebrafish retinal regeneration.

The adult zebrafish retina exhibits a robust regenerative response following light-induced photoreceptor cell death. This response is initiated by the Müller glia proliferating in the inner nuclear layer (INL), which gives rise to neuronal progenitor cells that continue to divide and migrate to the outer nuclear layer (ONL), where they differentiate into rod and cone photoreceptors. We previously conducted a microarray analysis of retinal gene expression at 16, 31, 51, 68, and 96 h of constant intense-light treatment to identify genes and their corresponding proteins that may be involved in the generation and proliferation of the neuronal progenitor cells.

The Tg(ccnb1:EGFP) transgenic zebrafish line labels proliferating cells during retinal development and regeneration.

Purpose: To create the Tg(ccnb1:EGFP)(nt18) zebrafish line that spatially and temporally labels retinal progenitor cells with enhanced green fluorescent protein (EGFP) during zebrafish retinal development and regeneration.

Methods: We cloned the 1.5 kb promoter region of the zebrafish cyclin B1 (ccnb1) gene upstream of the EGFP gene in the Tol2 vector, which was used to generate the stable Tg(ccnb1:EGFP)(nt18) transgenic zebrafish line.

Inhibition of Müller glial cell division blocks regeneration of the light-damaged zebrafish retina.

The adult zebrafish retina possesses a robust regenerative response. In the light-damaged retina, Müller glial cell divisions precede regeneration of rod and cone photoreceptors. Neuronal progenitors, which arise from the Müller glia, continue to divide and use the Müller glial cell processes to migrate to the outer nuclear layer and replace the lost photoreceptors.

Generation and characterization of transgenic zebrafish lines using different ubiquitous promoters.

Two commonly used promoters to ubiquitously express transgenes in zebrafish are the Xenopus laevis elongation factor 1 alpha promoter (XlEef1a1) and the zebrafish histone variant H2A.F/Z (h2afv) promoter. Recently, transgenes utilizing these promoters were shown to be silenced in certain adult tissues, particularly the central nervous system.

Time course analysis of gene expression during light-induced photoreceptor cell death and regeneration in albino zebrafish.

Constant intense light causes apoptosis of rod and cone photoreceptors in adult albino zebrafish. The photoreceptors subsequently regenerate from proliferating inner nuclear layer (INL) progenitor cells that migrate to the outer nuclear layer (ONL) and differentiate into rods and cones. To identify gene expression changes during this photoreceptor regeneration response, a microarray analysis was performed at five time points during the light treatment.

Retinal regional differences in photoreceptor cell death and regeneration in light-lesioned albino zebrafish.

Teleost fish regenerate retinal cells from a population of inner nuclear layer (INL) stem cells. To characterize photoreceptor regeneration in zebrafish (Danio rerio), adult albino fish were subjected to constant intense light to cause photoreceptor cell death. Retinal morphometry was performed on histological sections of control and light-lesioned albino retinas to compare the extent of light damage in the ventral, central and dorsal retinal regions.