scholarly journals Isolation and Characterization of Drosophila retinal degeneration B Suppressors

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 713-724 ◽  
Author(s):  
Don W Paetkau ◽  
Vecheslav A Elagin ◽  
Lisa M Sendi ◽  
David R Hyde
Keyword(s):  
Retinal Degeneration ◽  
Light Adaptation ◽  
Photoreceptor Cell ◽  
Light Response ◽  
Visual Transduction ◽  
Transfer Protein ◽  
Isolation And Characterization ◽  
Phosphatidylinositol Transfer ◽  
Normal Light

Abstract The Drosophila retinal degeneration B protein (RdgB) is a novel integral membrane phosphatidylinositol transfer protein required for photoreceptor cell viability and light response. We isolated one intragenic suppressor (rdgBsu100) and four autosomal suppressors of the hypomorphic rdgBKS222 retinal degeneration phenotype. The rdgBsu100 suppressor dramatically slowed rdgBKS222's photoreceptor degeneration without significantly improving the electroretinogram (ERG) light response. One autosomal recessive suppressor [su(rdgB)69] significantly slowed rdgBKS222 retinal degeneration and restored the ERG light response near to that of the wild type. Unlike all the previously characterized rdgB suppressors, the four new autosomal suppressors do not affect the ERG light response in rdgB+ flies. Only Su(rdgB)116 exhibited a mutant phenotype in a rdgB+ background, which was smaller R1-6 rhabdomeres. We also examined the extent to which two previously identified visual transduction mutations suppressed rdgB retinal degeneration. Absence of one of the light-activated calcium channels (trpCM) slowed the onset of rdgB-dependent degeneration. However, loss of protein kinase C (inaC209), which blocks photoreceptor cell deactivation, desensitization, and light adaptation, failed to suppress rdgB degeneration under normal light conditions. This demonstrates that TRP activity, but not INAC, is required for rapid rdgB-dependent degeneration.

scholarly journals The Phosphatidylinositol Transfer Protein Domain of Drosophila Retinal Degeneration B Protein Is Essential for Photoreceptor Cell Survival and Recovery from Light Stimulation

1997 ◽  
Vol 139 (2) ◽  
pp. 351-363 ◽  
Author(s):  
Scott C. Milligan ◽  
James G. Alb ◽  
Raya B. Elagina ◽  
Vytas A. Bankaitis ◽  
David R. Hyde
Keyword(s):  
Retinal Degeneration ◽  
Light Response ◽  
Protein Domain ◽  
Inherited Disease ◽  
Light Stimulation ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

The Drosophila retinal degeneration B (rdgB) gene encodes an integral membrane protein involved in phototransduction and prevention of retinal degeneration. RdgB represents a nonclassical phosphatidylinositol transfer protein (PITP) as all other known PITPs are soluble polypeptides. Our data demonstrate roles for RdgB in proper termination of the phototransduction light response and dark recovery of the photoreceptor cells. Expression of RdgB's PITP domain as a soluble protein (RdgB-PITP) in rdgB2 mutant flies is sufficient to completely restore the wild-type electrophysiological light response and prevent the degeneration. However, introduction of the T59E mutation, which does not affect RdgB-PITP's phosphatidylinositol (PI) and phosphatidycholine (PC) transfer in vitro, into the soluble (RdgB-PITP-T59E) or full-length (RdgB-T59E) proteins eliminated rescue of retinal degeneration in rdgB2 flies, while the light response was partially maintained. Substitution of the rat brain PITPα, a classical PI transfer protein, for RdgB's PITP domain (PITPα or PITPα-RdgB chimeric protein) neither restored the light response nor maintained retinal integrity when expressed in rdgB2 flies. Therefore, the complete repertoire of essential RdgB functions resides in RdgB's PITP domain, but other PITPs possessing PI and/or PC transfer activity in vitro cannot supplant RdgB function in vivo. Expression of either RdgB-T59E or PITPα-RdgB in rdgB+ flies produced a dominant retinal degeneration phenotype. Whereas RdgB-T59E functioned in a dominant manner to significantly reduce steady-state levels of rhodopsin, PITPα-RdgB was defective in the ability to recover from prolonged light stimulation and caused photoreceptor degeneration through an unknown mechanism. This in vivo analysis of PITP function in a metazoan system provides further insights into the links between PITP dysfunction and an inherited disease in a higher eukaryote.

scholarly journals Normal Light Response, Photoreceptor Integrity, and Rhodopsin Dephosphorylation in Mice Lacking Both Protein Phosphatases with EF Hands (PPEF-1 and PPEF-2)

2001 ◽  
Vol 21 (24) ◽  
pp. 8605-8614 ◽  
Author(s):  
Pradeep Ramulu ◽  
Matthew Kennedy ◽  
Wei-Hong Xiong ◽  
John Williams ◽  
Mitra Cowan ◽  
...  
Keyword(s):  
Retinal Degeneration ◽  
Light Response ◽  
Protein Product ◽  
Calcium Dependent ◽  
Cell Responses ◽  
Single Cell Responses ◽  
Rod Function ◽  
Low Levels ◽  
Ef Hands ◽  
Normal Light

ABSTRACT Rhodopsin dephosphorylation in Drosophila is a calcium-dependent process that appears to be catalyzed by the protein product of the rdgC gene. Two vertebrate rdgC homologs, PPEF-1 and PPEF-2, have been identified. PPEF-1 transcripts are present at low levels in the retina, while PPEF-2transcripts and PPEF-2 protein are abundant in photoreceptors. To determine if PPEF-2 alone or in combination with PPEF-1 plays a role in rhodopsin dephosphorylation and to determine if retinal degeneration accompanies mutation of PPEF-1 and/or PPEF-2, we have produced mice carrying targeted disruptions in thePPEF-1 and PPEF-2 genes. Loss of either or both PPEFs has little or no effect on rod function, as mice lacking both PPEF-1 and PPEF-2 show little or no changes in the electroretinogram and PPEF-2 −/− mice show normal single-cell responses to light in suction pipette recordings. Light-dependent rhodopsin phosphorylation and dephosphorylation are also normal or nearly normal as determined by (i) immunostaining ofPPEF-2 −/− retinas with the phosphorhodopsin-specific antibody RT-97 and (ii) mass spectrometry of C-terminal rhodopsin peptides from mice lacking both PPEF-1 and PPEF-2. Finally, PPEF-2 −/− retinas show normal histology at 1 year of age, and retinas from mice lacking both PPEF-1 and PPEF-2 show normal histology at 3 months of age, the latest time examined. These data indicate that, in contrast to loss of rdgC function in Drosophila, elimination of PPEF function does not cause retinal degeneration in vertebrates.

Characterization of mouse phosphatidylinositol transfer protein expressed in Escherichia coli

1994 ◽  
Vol 1213 (3) ◽  
pp. 309-318 ◽  
Author(s):  
Teunis B.H. Geijtenbeek ◽  
Ellen de Groot ◽  
Jürgen van Baal ◽  
Freek Brunink ◽  
Jan Westerman ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

Expression and characterization of a barley phosphatidylinositol transfer protein structurally homologous to the yeast Sec14p protein

Plant Science ◽  
2016 ◽  
Vol 246 ◽  
pp. 98-111 ◽  
Author(s):  
Agnieszka Kiełbowicz-Matuk ◽  
Ewa Banachowicz ◽  
Anna Turska-Tarska ◽  
Pascal Rey ◽  
Tadeusz Rorat
Keyword(s):  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Phosphatidylinositol Transfer

Light-induced retinal degeneration in rdgB (retinal degeneration B) mutant of Drosophila: Electrophysiological and morphological manifestations of degeneration

1989 ◽  
Vol 2 (6) ◽  
pp. 529-539 ◽  
Author(s):  
Chaim T. Rubinstein ◽  
Shoshana Bar-Nachum ◽  
Zvi Selinger ◽  
Baruch Minke
Keyword(s):  
Retinal Degeneration ◽  
Early Stage ◽  
Light And Electron Microscopy ◽  
Bright Light ◽  
Light Response ◽  
Photoreceptor Cells ◽  
Dark Cycle ◽  
Voltage Dependent ◽  
Normal Light ◽  
Two Phases

AbstractQuantitative light and electron microscopy was used to monitor the extent of retinal degeneration as a function of age and temperature in the white-eyed rdgBKS222 mutant of Drosophila melanogaster. Parallel measurements of the electroretinogram (ERG) of the degenerating retina reveal a new phenomenon – the appearance of spike potentials following illumination with bright light. These spikes, which do not appear in the normal fly retina, have a relatively long duration (20–50 ms), regenerative properties, and a rate of occurrence which increases with increasing light intensity. The spikes differed from the light response in being more susceptible to CO2 and to cuts in the eye. The spikes completely disappeared at low extracellular Ca2+ levels which did not reduce the amplitude of the light response. The spike potentials become triphasic when the recording electrode is advanced to the level of the basement membrane. This suggests that the spike potentials originate from the photoreceptor axons as a result of synchronous opening of voltage-dependent channels in a large number of photoreceptor cells. The occurrence of spike potentials during the process of degeneration was studied. Two phases can be distinguished: (1) Spike potentials appear in retinae of 2–3-day-old flies which display few morphological signs of degeneration. The frequency of appearance of spike potentials decreases in retinae of 14–16-day-old flies which show extensive degeneration of the R1–6 photoreceptor cells but no degeneration of the central R7,8 cells. (2) Spike potentials appear more frequently again in flies of 22–24 d of age. This is probably a consequence of degeneration of the remaining R7,8 photoreceptor cells. Temperature and the light-dark cycle had a critical effect on degeneration. Eight-day-old mutants raised at 19°C in a normal light-dark cycle showed only little degeneration. Eight-day-old mutants raised at 24°C showed only a slight degeneration when raised in the dark. However, the degree of degeneration was greatly enhanced in the mutants raised at 24°C under a light-dark cycle regime.The combined electrophysiological and morphological study of the degeneration, as a function of age and temperature, revealed that (1) the degeneration process takes place even in darkness, but at a slow rate, while light greatly accelerates the degeneration. (2) The degeneration is negligible at 19°C, even during light, in the first week after eclosion. (3) The appearance of spike potentials at an early stage of the degeneration suggests that changes in the plasma membrane of the photoreceptor cells manifest at an initial stage of the degeneration process.

scholarly journals Localization of Drosophila retinal degeneration B, a membrane-associated phosphatidylinositol transfer protein

1993 ◽  
Vol 122 (5) ◽  
pp. 1013-1022 ◽  
Author(s):  
TS Vihtelic ◽  
M Goebl ◽  
S Milligan ◽  
JE O'Tousa ◽  
DR Hyde
Keyword(s):  
Retinal Degeneration ◽  
Amino Terminal ◽  
Transfer Protein ◽  
Phosphatidylinositol Transfer Protein ◽  
Protein Functions ◽  
Drosophila Brain ◽  
Phosphatidylinositol Transfer ◽  
Acid Domain ◽  
Membrane Spanning

The Drosophila retinal degeneration B (rdgB) mutation causes abnormal photoreceptor response and light-enhanced retinal degeneration. Immunoblots using polyclonal anti-rdgB serum showed that rdgB is a 160-kD membrane protein. The antiserum localized the rdgB protein in photoreceptors, antennae, and regions of the Drosophila brain, indicating that the rdgB protein functions in many sensory and neuronal cells. In photoreceptors, the protein localized adjacent to the rhabdomeres, in the vicinity of the subrhabdomeric cisternae. The rdgB protein's amino-terminal 281 residues are > 40% identical to the rat brain phosphatidylinositol transfer protein (PI-TP). A truncated rdgB protein, which contains only this amino-terminal domain, possesses a phosphatidylinositol transfer activity in vitro. The remaining 773 carboxyl terminal amino acids have additional functional domains. Nitrocellulose overlay experiments reveal that an acidic amino acid domain, adjacent to the PI transfer domain, binds 45Ca+2. Six hydrophobic segments are found in the middle of the putative translation product and likely function as membrane spanning domains. These results suggest that the rdgB protein, unlike the small soluble PI-TPs, is a membrane-associated PI-TP, which may be directly regulated by light-induced changes in intracellular calcium.

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