ATG8-Binding UIM Proteins Define a New Class of Autophagy Adaptors and Receptors
Authors: Zhihua Hua
Richard D Vierstra
Fionn McLoughlin
Sujina Mali
Richard S Marshall
Journal: Cell (2019)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: AIM; ATG8; ATG8-interacting motif; Arabidopsis; CDC48/p97
Affiliations: Department of Biology, Washington University in St. Louis, St. Louis, MO 63130, USA., Department of Environmental and Plant Biology, Ohio University, Athens, OH 45701, USA., Department of Biology, Washington University in St. Louis, St. Louis, MO 63130, USA. Electronic address: .

Abstract:  During autophagy, vesicle dynamics and cargo recruitment are driven by numerous adaptors and receptors that become tethered to the phagophore through interactions with lipidated ATG8/LC3 decorating the expanding membrane. Most currently described ATG8-binding proteins exploit a well-defined ATG8-interacting motif (AIM, or LC3-interacting region [LIR]) that contacts a hydrophobic patch on ATG8 known as the LIR/AIM docking site (LDS). Here we describe a new class of ATG8 interactors that exploit ubiquitin-interacting motif (UIM)-like sequences for high-affinity binding to an alternative ATG8 interaction site. Assays with candidate UIM-containing proteins together with unbiased screens identified a large collection of UIM-based ATG8 interactors in plants, yeast, and humans. Analysis of a subset also harboring ubiquitin regulatory X (UBX) domains revealed a role for UIM-directed autophagy in clearing non-functional CDC48/p97 complexes, including some impaired in human disease. With this new class of adaptors and receptors, we greatly extend the reach of selective autophagy and identify new factors regulating autophagic vesicle dynamics. 

Retracted: No

PubPeer comments:
 1. Figure 2A-C: No background at all for negative controls results? This means that no yeast was dropped onto the corresponding place on the plates. Same for Figure 2G, Figure S2C. ![image] ![image] This is obvious when comparing to a normal yeast two hybrid figure for example other Y2H in the same paper like Figure 1A-B, which shows clearly dead yeast dot background. Demonstration of a normal background for yeast two hybrid assay: ![image] , 
 2. Figure 2F: The three micrographs highlighted are black boxes with no background signals at all. Normal background shall be displayed as the rest, showing clear background noise signals after curve adjustment. ![image] , 
 3. Figure 7A and 7D: The two u201cH3u201d blots as controls are the same blot. Please see the crop from the original figures and after curve adjustment with direct side by side comparison. ![image] Side by side comparison: (upper panel is H3 blot from 7A, lower panel is H3 blot from 7D.) ![image] , 
 4. Figure S1: The fluorescence images from Figure S1 D lower panel are purely duplicates from the upper panel except the two in the centre. Please see the curve adjusted figures for clearer comparison. ![image] ![image] , 
 7-14-2021 Dear PubPeer: Thanks for the inquiry related to the Marshall et al. (2019) Cell publication about possible inconsistencies irregularities about some of its figures. Like you, we the authors are deeply committed to publishing accurate scientific data and have taken your notification seriously. Fortunately, we have found all the original results highlighted by you as possibly being problematic and are now working with Cell to fix any errors. We have also gone over all the other figures to confirm accuracy. From our perspective, it looks like some of the figures highlighted by PubPeer are actually correct and that some have been published in error. Tracing back the errors, it appears that our originally formatted figures were correct in all cases but then some of the panels became altered substituted by various authors during the iterative improvements rearrangements of the figures before final publication. Our time-stamps on the succession of figure files allowed us to see when the changes were introduced in error. It appears to be a combined set of errors involving several authors, with none of the authors detecting the problems in the final product. Had we seen these errors, we would have without question fixed them promptly. Importantly, these placement errors in no way altered the outcomes nor our interpretations in the Cell publication, and can easily be rectified by replacing the panels published in error. We hope to have the corrections complete in the near future. Sincerely, Richard D. Vierstra Richard S. Marshall , 
 9-29-2021 Dear Colleagues in PubPeer, We, like you, are committed to publishing accurate and meaningful scientific data and became concerned about possible errors in this publication upon receiving the above notification from PubPeer. We have now gone to great lengths to research the outlined concerns and associated raw data, and when and where any errors were introduced. We affirm that the conclusions of the paper are correct, and notably in some cases have since been confirmed by others. However, YES we did find some errors, which either involved: (i) incorrect placement of immunoblots using antibodies against histone H3 that we used routinely as a control for protein loading (Figure 7D), and (ii) incorrect placement of control BiFC images (Figure S1D). Fortunately, however, not all possible errors implied by PubPeer were correct. The Y2H images in Figures 2A, 2B, 2C, 2G and S2C contain the correct data, while the BiFC panels in Figure 2F include the correct images. Below we provide information regarding the images that we feel to be correct; an official correction will be published in Cell in due course to address the remaining errors. Figures 2A, 2B, 2C, 2G and S2C For the indicated Y2H panels implied as incorrect, we show the published data, original TIF scans, and brightness enhanced TIF scans for each Y2H plate on pages 1 and 2 of the attached files (to follow in subsequent comments). As you will hopefully see from the original scans upon brightening, yeast cells were indeed spotted in each position of the plate. However, it does appear that during preparation of the Figure, which involved conversion of these RGB images generated by a laser scanner to grayscale and then adjustment of the brightness and contrast of the images, we lost view of the spotted yeast that failed to grow colonies. Despite this, the Y2H experiments were conducted correctly and the results were properly interpreted. Cell have requested that we post this information here, rather than include it in a formal correction. Figure 2F Regarding the concerns related to the BiFC data on Figure 2F, on pages 3 and 4 of the attached files (see subsequent comments), we show the PUX7 and PUX13 BiFC images as published, along with the original TIF files directly saved from the microscope (bright field bottom left, YFP signal top right, and merged images top left), plus versions of these with enhanced brightness. We hope that in all three dark panels of concern, the viewer will observe that, upon increasing the brightness, background signal is indeed visible in the analysis of both PUX7 and PUX13 (for example in several panels, you can see the outline of guard cells). Given that all the images were taken with identical microscope settings, it is likely the case that the background signal was particularly low in these samples, which can sometimes occur depending on the age of the infiltrated leaf, or the focal plane that is used. We can therefore find no errors in this Figure or its conclusions. As stated above, remaining errors will be corrected in an upcoming issue of Cell in the near future. Yours sincerely, Prof. Richard D. Vierstra and Dr. Richard S. Marshall. , 
 ![file] ![file] ![file] ![file] , 
 I am not sure about the author-provided originals showing the yeast assays. Both the published figures as well as the provided originals look very regular, without the small irregularities in shape and position normally observed in these assays. They look like nearly-perfectly white circles on a perfectly black background. In addition, in the few cases where some additional features can be observed in the published photos, these appear to be missing from the original scans provided above by the authors. In Figure 2C, the published photo shows two small white dots, if you make the image a bit brighter (marked with green arrows below). These should also be visible on the original scan-with increased brightness, but I cannot find them. Published Figure 2C: ![file] Original scan for Figure 2C: ![file] Similarly, the published Supplemental Figure S2C shows small white dots in the background (green arrows) and irregularities in some of the yeast dots (orange arrows) that appear to be missing from the original scans provided by the author. ![file] , 
 Figure 6ABD. * Some panels here appear to show duplicated areas, which I marked with boxes of the same color. ![file] , 
 Figure 6F, 6N. * Some panels here appear to show duplicated areas, which I marked with boxes of the same color. ![file] , 
 Supplemental Figure S7G. * The top panel appears to show duplicated areas, marked with green boxes. ![file] , 
 As pointed out on Twitter by @Hallfiry, there are two duplicated panel in the [PubMedCentral version] of this paper, which are not present in the [Cell Press version] of this paper. The duplications are shown below with red and cyan boxes. One of the duplications involves a 180 degrees rotation. Could the authors please clarify if why these duplications have been removed from the Cell Press version? The PMC version was published after the Cell Press version, but perhaps the Cell Press version has been silently updated since? Was there a corrigendum associated with this? The March 2022 Corrigendum only mentions new versions of Figures 7D and S1D, but maybe I am overlooking something . ![file] , 
 Dear PubPeer Commentors: I apologize for the various errors inconsistencies in this paper. The lead author, Dr. RIchard Marshall is no longer in the lab has not yet responded to my requests for comment related to several of your recent concerns. Prior PubPeer concerns were addressed by us through the Editor of Cell, for which we published a Correction as requested by the journal. Related to the odd differences in the version of Figure 7BE available in PubMed Central as compared to that published by Cell, I do not understand how these arose but can see the issue when downloading both versions. As far as I am aware, Cell has not silently updated the figures. The final PDF of the paper available in Cell also matches my PDF of the paper download in May 2,2019. My files of the manuscript also have the final version of Figure 7 generated in February 13, 2019; it is identical to that published by Cell. It is attached. However, I have found an older version of Figure 7 that has the duplicated panels in Figure 7BE. How these erroneously ended up in the PubMed Central version is unknown to me. If you have advice and how to repair this error, please let me know. Despite the irregularities, I am not aware that any of the conclusions from the paper have been compromised. If I find otherwise, I will let you know. Sorry for the confusion. ![file] Rick Vierstra ,