UIUC Plant Physiology JC (2019/08/12): α-carboxysome formation
The UIUC Plant Physiology journal club reviewed the preprint “α-carboxysome formation is mediated by the multivalent and disordered protein CsoS2
” (doi: https://doi.org/10.1101/708164
) by Oltrogge et al. 2019. The paper describes the biochemical characterization of the CsoS2 protein involved in carboxysome assembly, identifying a repeat peptide region that makes weak electrostatic interactions with rubisco through the use of bio-layer interferometry (BLI) and x-ray crystallography. The authors identified evolutionary conserved residues through protein sequence comparisons and the sites of interaction between these residues and rubisco through protein X-ray crystallography.
We found the paper to be very well written, well presented and valuable addition to knowledge about alpha carboxysome assembly. Our journal club assessed the paper as part of a learning exercise about how to make work accessible to a wide audience.
Participants first learned about the “and-but-therefore (ABT)” model of paper writing popularized by Randy Olsen in his freely available book "Huston, we have a narrative
", that can be used throughout the manuscript to help maintain the reader’s interest. Focusing on the abstract we found it to contain many of aspects of the ABT model. We also thought it could potentially be strengthened by including a stronger “but” phrase which generally represents the question under consideration. It was suggested that this phrase would start with the fact that there is little knowledge about how the carboxysome is assembled, and some members of the club questioned if the ongoing carboxysome engineering efforts might be mentioned as relevant to the wider importance of the work (either in the abstract or the discussion).
One aspect we found particularly interesting was the similarities between CsoS2 and the algal protein EPYC which has been implicated in aggregation of rubisco in the pyrenoid. These appeared to us to an important point, and the reason to include information about CsoS2 as an intrinsically disordered protein (IDP) that could perhaps be emphasized more. As we were not familiar with the PONDR-FIT disorder score, we would have found it helpful to have a little more explanation as to its importance and interpretation. Overall we liked the approach for analyzing IDPs and thought it was an impressive effort to successfully crystallize the CsoS2 peptide with rubisco.
In addition, we assessed the presentation of figures, we particularly liked the use of consistent colouring throughout, the choice of clearly legible font sizes on all graphs and the helpful diagrams to illustrate biochemical procedures, such as the BLI procedure in Fig 2b. One consideration is whether the choice of colors is colorblind friendly, using the app color oracle, several of the colors are indistinguishable in all the figures analysed. We also thought inclusion of legend titles would help guide readers on how best to interpret the data.
We thought the X-ray crystallography data was presented in a clear and helpful manner, displaying what the individual residue interactions were between the bpeptide and rubisco. If it could be improved further it may be by inclusion of a label of rubisco for non-experts who may not immediately associate CbbL and CbbS as subunits. Finally, we particularly liked Figure 5 as it neatly summarized the proposed role of CsoS2 in carboxysome assembly.
Other thoughts included:
- It would be interesting to include discussion of why the full length CsoS2 peptide does not appear to bind rubisco.
- The paper tied up loose ends and did a good job of using multiple approaches to build evidence for the direct interaction of CsoS2 and rubisco.