Retraction notice to "Peptide-induced formation of protein aggregates and amyloid fibrils in human and Guinea pig αA-crystallins under physiological conditions of temperature and pH" [Exp. Eye Res. 179 (2019) 193-205].

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the authors. The senior author contacted the journal in a forthright manner, in an effort to preserve the scientific integrity of the literature, after discovering a significant error in the results reported in the article. The authors were recently made aware of a paper by Kim et al. (Nature Commun. 2019) which shows a spirosome structure (the enzyme aldehyde-alcohol dehydrogenase) present in E. coli (Fig. 5a) that is very similar to the structure the authors thought formed when synthetic alpha A crystallin (66-80) peptide was incubated for 24 h with recombinant guinea pig alpha A insert crystallin (see Kumarasamy et al. Figs. 7C and F, and Fig. 9). Subsequent to publication of their report, the authors later found a number of images that showed what appeared to be the same structure present in samples of their presumably purified recombinant guinea pig alpha A insert crystallin which had been incubated without peptide for 24 h. Hence, the authors now conclude that the structures shown in Figs. 7C and F, and Fig. 9 of their article published in this journal are actually due to E. coli contaminant aldehyde-alcohol dehydrogenase. The authors deeply regret this error and any inconvenience it may have caused.

Adipocyte lipolysis affects Perilipin 5 and cristae organization at the cardiac lipid droplet-mitochondrial interface.

This study investigated the effects of elevated fatty acid (FA) supply from adipose tissue on the ultrastructure of cardiac lipid droplets (LDs) and the expression and organization of LD scaffold proteins perilipin-2 (PLIN2) and perilipin-5 (PLIN5). Stimulation of adipocyte lipolysis by fasting (24 h) or ß3-adrenergic receptor activation by CL316, 243 (CL) increased cardiac triacylglycerol (TAG) levels and LD size, whereas CL treatment also increased LD number. LDs were tightly associated with mitochondria, which was maintained during LD expansion. Electron tomography (ET) studies revealed continuity of LD and smooth endoplasmic reticulum (SER), suggesting interconnections among LDs. Under fed ad libitum conditions, the cristae of mitochondria that apposed LD were mostly organized perpendicularly to the tangent of the LD surface. Fasting significantly reduced, whereas CL treatment greatly increased, the perpendicular alignment of mitochondrial cristae. Fasting and CL treatment strongly upregulated PLIN5 protein and PLIN2 to a lesser extent. Immunofluorescence and immuno-electron microscopy demonstrated strong targeting of PLIN5 to the cardiac LD-mitochondrial interface, but not to the mitochondrial matrix. CL treatment augmented PLIN5 targeting to the LD-mitochondrial interface, whereas PLIN2 was not significantly affected. Together, our results support the concept that the interface between LD and cardiac mitochondria represents an organized and dynamic "metabolic synapse" that is highly responsive to FA trafficking.

RETRACTED: Peptide-induced formation of protein aggregates and amyloid fibrils in human and guinea pig αA-crystallins under physiological conditions of temperature and pH.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the authors. The senior author contacted the journal in a forthright manner, in an effort to preserve the scientific integrity of the literature, after discovering a significant error in the results reported in the article. The authors were recently made aware of a paper by Kim et al. (Nature Commun. 2019) which shows a spirosome structure (the enzyme aldehyde-alcohol dehydrogenase) present in E. coli (Fig. 5a) that is very similar to the structure the authors thought formed when synthetic alpha A crystallin (66-80) peptide was incubated for 24 h with recombinant guinea pig alpha A insert crystallin (see Kumarasamy et al., Figs. 7C and F, and Fig. 9). Subsequent to publication of their report, the authors later found a number of images that showed what appeared to be the same structure present in samples of their presumably purified recombinant guinea pig alpha A insert crystallin which had been incubated without peptide for 24 h. Hence, the authors now conclude that the structures shown in Figs. 7C and F, and Fig. 9 of their article published in this journal are actually due to E. coli contaminant aldehyde-alcohol dehydrogenase. The authors deeply regret this error and any inconvenience it may have caused.

ARL2BP, a protein linked to retinitis pigmentosa, is needed for normal photoreceptor cilia doublets and outer segment structure.

The outer segment (OS) of photoreceptor cells is an elaboration of a primary cilium with organized stacks of membranous disks that contain the proteins needed for phototransduction and vision. Though ciliary formation and function has been well characterized, little is known about the role of cilia in the development of photoreceptor OS. Nevertheless, progress has been made by studying mutations in ciliary proteins, which often result in malformed OSs and lead to blinding diseases. To investigate how ciliary proteins contribute to OS formation, we generated a knockout (KO) mouse model for ARL2BP, a ciliary protein linked to retinitis pigmentosa. The KO mice display an early and progressive reduction in visual response. Before photoreceptor degeneration, we observed disorganization of the photoreceptor OS, with vertically aligned disks and shortened axonemes. Interestingly, ciliary doublet microtubule (MT) structure was also impaired, displaying open B-tubule doublets, paired with loss of singlet MTs. On the basis of results from this study, we conclude that ARL2BP is necessary for photoreceptor ciliary doublet formation and axoneme elongation, which is required for OS morphogenesis and vision.

Vacuolar protein sorting 13C is a novel lipid droplet protein that inhibits lipolysis in brown adipocytes.

OBJECTIVE: Brown adipose tissue (BAT) thermogenesis depends on the mobilization and oxidation of fatty acids from intracellular lipid droplets (LD) within brown adipocytes (BAs); however, the identity and function of LD proteins that control BAT lipolysis remain incomplete. Proteomic analysis of mouse BAT subcellular fractions identified vacuolar protein sorting 13C (VPS13C) as a novel LD protein. The aim of this work was to investigate the role of VPS13C on BA LDs. METHODS: Biochemical fractionation and high resolution confocal and immuno-transmission electron microscopy (TEM) were used to determine the subcellular distribution of VPS13C in mouse BAT, white adipose tissue, and BA cell culture. Lentivirus-delivered shRNA was used to determine the role of VPS13C in regulating lipolysis and gene expression in cultured BA cells. RESULTS: We found that VPS13C is highly expressed in mouse BAT where it is targeted to multilocular LDs in a subspherical subdomain. In inguinal white adipocytes, VPS13C was mainly observed on small LDs and ß3-adrenergic stimulation increased VPS13C in this depot. Silencing of VPS13C in cultured BAs decreased LD size and triglyceride content, increased basal free fatty acid release, augmented the expression of thermogenic genes, and enhanced the lipolytic potency and efficacy of isoproterenol. Mechanistically, we found that BA lipolysis required activation of adipose tissue triglyceride lipase (ATGL) and that loss of VPS13C greatly increased the association of ATGL to LDs. CONCLUSIONS: VPS13C is present on BA LDs where is targeted to a distinct subdomain. VPS13C limits the access of ATGL to LD and loss of VPS13C elevates lipolysis and promotes oxidative gene expression.

An inducible amphipathic helix within the intrinsically disordered C terminus can participate in membrane curvature generation by peripherin-2/rds.

Peripherin-2/rds is required for biogenesis of vertebrate photoreceptor outer segment organelles. Its localization at the high-curvature rim domains of outer segment disk membranes suggests that it may act to shape these structures; however, the molecular function of this protein is not yet resolved. Here, we apply biochemical, biophysical, and imaging techniques to elucidate the role(s) played by the protein's intrinsically disordered C-terminal domain and an incipient amphipathic α-helix contained within it. We investigated a deletion mutant lacking only this α-helix in stable cell lines and Xenopus laevis photoreceptors. We also studied a soluble form of the full-length ∼7-kDa cytoplasmic C terminus in cultured cells and purified from Escherichia coli The α-helical motif was not required for protein biosynthesis, tetrameric subunit assembly, tetramer polymerization, localization at disk rims, interaction with GARP2, or the generation of membrane curvature. Interestingly, however, loss of the helical motif up-regulated membrane curvature generation in cellulo, introducing the possibility that it may regulate this activity in photoreceptors. Furthermore, the incipient α-helix (within the purified soluble C terminus) partitioned into membranes only when its acidic residues were neutralized by protonation. This suggests that within the context of full-length peripherin-2/rds, partitioning would most likely occur at a bilayer interfacial region, potentially adjacent to the protein's transmembrane domains. In sum, this study significantly strengthens the evidence that peripherin-2/rds functions directly to shape the high-curvature rim domains of the outer segment disk and suggests that the protein's C terminus may modulate membrane curvature-generating activity present in other protein domains.

Effect of a blueberry nutritional supplement on macronutrients, food group intake, and plasma vitamin E and vitamin C in US athletes.

Antioxidants from a blueberry beverage may impact plasma vitamins. We examined vitamins/food selection in 12 college athletes during 30 days compared with placebo. Blood was collected before and after exercise at the beginning of the study (day 1) and then after a 30-day period of taking a daily supplemental beverage (day 30). The six trials involved blood that was drawn pre-beverage ingestion/pre-exercise (trials 1 and 4), post-beverage ingestion/pre-exercise (trials 2 and 5), and post-beverage ingestion/1 h post-exercise (trials 3 and 6), on day 1 (trials 1, 2, and 3) and day 30 (trials 4, 5, and 6). Analysis of variance revealed non-significant differences for macronutrient or gamma-tocopherol and vitamin C intakes by food frequency questionnaire or plasma vitamins by liquid chromatography. There was a trend (P = 0.083) in the group x time interaction for alpha-tocopherol intake by repeated-measures analysis of variance. Blueberry alpha-tocopherol (23.91 +/- 9.31 mg) was significantly (P < 0.05) higher than placebo alpha-tocopherol intake (7.59 +/- 0.95 mg) on day 1, but not on day 30 (blueberry, alpha-tocopherol = 9.04 +/- 2.35 mg, placebo, alpha-tocopherol = 11.46 +/- 3.65 mg) by pairwise comparisons. Blueberry supplementation did not affect plasma vitamin concentrations or gamma-tocopherol and vitamin C intakes, and may reduce alpha-tocopherol intake in those starting with a higher alpha-tocopherol intake, yet not altering athletes' eating habits.

Characterization of melanophore morphology by fractal dimension analysis.

Fractal or focal dimension (FD) analysis is a valuable tool to identify physiologic stimuli at the cellular and tissue levels that allows for quantification of cell perimeter complexity. The FD analysis was determined on fluorescence images of caffeine- or epinephrine-treated (or untreated control) killifish Fundulus heteroclitus (Linneaus) melanophores in culture. Cell perimeters were indicated by rhodamine-phalloidin labeling of cortical microfilaments using box-counting FD analysis. Caffeine-treated melanophores displayed dispersed melanosomes in cells with less serrated edges and reduced FD and complexity. Complexity in epinephrine-treated cells was significantly higher than the caffeine-treated cells or in the control. Cytoarchitectural variability of the cell perimeter is expected because cells change shape when cued with agents. Epinephrine-treated melanophores demonstrated aggregated melanosomes in cells with more serrated edges, significantly higher FD and thus complexity. Melanophores not treated with caffeine or epinephrine produced variable distributions of melanosomes and resulted in cells with variably serrated edges and intermediate FD with a larger SE of the regression and greater range of complexity. Dispersion of melanosomes occurs with rearrangements of the cytoskeleton to accommodate centrifugal distribution of melanosomes throughout the cell and to the periphery. The loading of melanosomes onto cortical microfilaments may provide a less complex cell contour, with the even distribution of the cytoskeleton and melanosomes. Aggregation of melanosomes occurs with rearrangements of the cytoskeleton to accommodate centripetal distribution of melanosomes. The aggregation of melanosomes may contribute to centripetal retraction of the cytoskeleton and plasma membrane. The FD analysis is, therefore, a convenient method to measure contrasting morphologic changes within stimulated cells.

Morphological studies on the mechanisms of pigmentary organelle transport in fish xanthophores and melanophores.

Pigmentary organelle translocations within fish chromatophores undergo physiological color changes when exposed to external signals. Chromatophores can be isolated in high yields, and their pigmentary organelles can be tracked readily by microscopy. The combined efforts of morphology and biomolecular chemistry have led to the identification of and determination of the interrelationships between cytoskeletal elements and accessory proteins, motor molecules, cytomatrix, and pigmentary organelles of various sizes. Fish chromatophores have been classified as fast, intermediate, and slow translocators, based on the relative numbers of microtubules. Studies on cultured goldfish (Carassius auratus L.) xanthophores for over 20 years have demonstrated that in this slow translocator, tubulovesicular structures of the smooth endoplasmic reticular (SER) cisternae are involved in the disperson and aggregation of associated carotenoid droplets (CD) with some involvement of cytoskeletal elements. Killifish (Fundulus heteroclitus L.) melanophore, a fast translocator, was also examined. Recent work demonstrates a bright fluorescent "starburst"-like spot that we call an actin filament-organizing center (AFOC) with radiating microfilaments, akin to the microtubule-organizing center (MTOC) with radiating microtubules. Melanosomes translocate single-file on microtubules and are not associated with SER cisternae. Slower CD dispersion or aggregation in goldfish xanthophores seems to be predominantly microfilament-based transport, or microfilament- and microtubule-based transport, respectively. Faster melanosome translocations in killifish melanophores are based on microtubules, with our evidence indicating microfilament involvement. Neural crest-derived chromatophores are models for vesicular transport in axons, and immunocytochemical and imaging technologies may help to elucidate the cellular transport mechanisms.

Morphological studies on microfilaments and their organizing center in killifish (Fundulus heteroclitus L.) melanophores.

Fish chromatophores serve as excellent study models for cytoskeleton-dependent organelle translocations because the distribution of pigmentary organelles can be observed against a time frame by microscopy. In this study the distribution of microfilaments along with microtubules in cultured melanophores of the killifish (Fundulus heteroclitus Linneaus) are examined using whole-cell transmission electron microscopy (WCTEM), fluorescence, and laser scanning confocal microscopy. Dispersing, dispersed, aggregating and aggregated states of pigment are induced by adding either caffeine (for dispersion) or epinephrine (for aggregation) to the cells in a standard culture medium. The cells that exhibited a random melanosome distribution in the standard culture media without these two reagents, served as the control. The results indicate that: (i) a structure considered to be the actin-filament organizing center (AFOC) is in close proximity to the microtubule-organizing center (MTOC); (ii) the radial layout of microfilaments remains similar over four physiological states of pigmentary response with the exception of epinephrine-aggregated pigment, in which the aggregate blocks the viewing of the AFOC and central microfilament rays, yet radial microfilaments, whether central and/or peripheral, are apparent in all physiological states of distribution; and (iii) microfilaments serve, together with microtubules, as scaffolding for melanosomes which migrate in bi-directional rows on cross-bridges, thus shedding light on the mechanisms for orderly melanosome translocations in a structural continuum.