Prevalence and species distribution of the low-complexity, amyloid-like, reversible, kinked segment structural motif in amyloid-like fibrils

Abstract

Membraneless organelles (MLOs) are vital and dynamic reaction centers in cells that compartmentalize the cytoplasm in the absence of a membrane. Multivalent interactions between protein low-complexity domains contribute to MLO organization. Previously, we used computational methods to identify structural motifs termed low-complexity amyloid-like reversible kinked segments (LARKS) that promote phase transition to form hydrogels and that are common in human proteins that participate in MLOs. Here, we searched for LARKS in the proteomes of six model organisms: Homo sapiens, Drosophila melanogaster, Plasmodium falciparum, Saccharomyces cerevisiae, Mycobacterium tuberculosis, and Escherichia coli to gain an understanding of the distribution of LARKS in the proteomes of various species. We found that LARKS are abundant in M. tuberculosis, D. melanogaster, and H. sapiens but not in S. cerevisiae or P. falciparum. LARKS have high glycine content, which enables kinks to form as exemplified by the known LARKS-rich amyloidogenic structures of TDP43, FUS, and hnRNPA2, three proteins that are known to participate in MLOs. These results support the idea of LARKS as an evolved structural motif. Based on these results, we also established the LARKSdb Web server, which permits users to search for LARKS in their protein sequences of interest. Membraneless organelles (MLOs) are vital and dynamic reaction centers in cells that compartmentalize the cytoplasm in the absence of a membrane. Multivalent interactions between protein low-complexity domains contribute to MLO organization. Previously, we used computational methods to identify structural motifs termed low-complexity amyloid-like reversible kinked segments (LARKS) that promote phase transition to form hydrogels and that are common in human proteins that participate in MLOs. Here, we searched for LARKS in the proteomes of six model organisms: Homo sapiens, Drosophila melanogaster, Plasmodium falciparum, Saccharomyces cerevisiae, Mycobacterium tuberculosis, and Escherichia coli to gain an understanding of the distribution of LARKS in the proteomes of various species. We found that LARKS are abundant in M. tuberculosis, D. melanogaster, and H. sapiens but not in S. cerevisiae or P. falciparum. LARKS have high glycine content, which enables kinks to form as exemplified by the known LARKS-rich amyloidogenic structures of TDP43, FUS, and hnRNPA2, three proteins that are known to participate in MLOs. These results support the idea of LARKS as an evolved structural motif. Based on these results, we also established the LARKSdb Web server, which permits users to search for LARKS in their protein sequences of interest. A new area of cell biology is the study of membraneless organelles (MLOs) in the organization of cellular structures and metabolism. Many MLOs are RNA and protein assemblies that fulfill specific functions for the cell. Examples of MLOs in human cells include P bodies that degrade mRNA, stress granules (SGs) that store mRNA during stresses, and the nucleolus that processes rRNA. MLOs also function in other organisms such as germline P granules in Caenorhabditis elegans and SGs in yeast. The aforementioned organelles are not enveloped by membranes to partition them from the cytoplasm but instead organize through multivalent networks of homotypic and heterotypic reversible interactions between proteins and nucleic acids (1Alberti S. Gladfelter A. Mittag T. Considerations and challenges in studying liquid-liquid phase separation and biomolecular condensates.Cell. 2019; 176: 419-434Abstract Full Text Full Text PDF PubMed Scopus (681) Google Scholar, 2Li P. Banjade S. Cheng H.-C. Kim S. Chen B. Guo L. Llaguno M. Hollingsworth J.V. King D.S. Banani S.F. Russo P.S. Jiang Q.-X. Nixon B.T. Rosen M.K. Phase transitions in the assembly of multi-valent signaling proteins.Nature. 2012; 483: 336-340Crossref PubMed Scopus (1103) Google Scholar). These reversible networks allow MLOs to be dynamic. They may assemble and disassemble in response to stimuli like SGs do in response to stresses and dissolve as stresses subside. Proteins in MLOs often contain low-complexity domains (LCDs) that help to drive reversible organization (3Kato M. McKnight S.L. A solid-state conceptualization of information transfer from gene to message to protein.Annu. Rev. Biochem. 2018; 87: 351-390Crossref PubMed Scopus (62) Google Scholar, 4Kato M. Han T.W. Xie S. Shi K. Du X. Wu L.C. Mirzaei H. Goldsmith E.J. Longgood J. Pei J. Grishin N.V. Frantz D.E. Schneider J.W. Chen S. Li L. et al.Cell-free formation of RNA granules: Low complexity sequence domains form dynamic fibers within hydrogels.Cell. 2012; 149: 753-767Abstract Full Text Full Text PDF PubMed Scopus (1183) Google Scholar). LCDs are regions of proteins with significant biases for one or a few amino acids. An example is the LCD of FUS where four amino acids glycine, tyrosine, serine, and glutamine account for 80% of the LCD composition. The LCD from FUS has also been termed an intrinsically disordered region (IDR) or a prion-like domain. The LCD of FUS is an IDR because for most of the time it lacks a defined globular structure. In fact, LCDs are a reasonable proxy for IDRs (5Wootton J.C. Non-globular domains in protein sequences: Automated segmentation using complexity measures.Comput. Chem. 1994; 18: 269-285Crossref PubMed Scopus (376) Google Scholar), but this is not always the case (e.g., collagen proteins that are low in sequence complexity but have a defined structure). Prion-like domains technically refer to a domain that resembles a yeast prion sequence that is rich in asparagine and glutamine (6Cascarina S.M. Paul K.R. Ross E.D. Manipulating the aggregation activity of human prion-like proteins.Prion. 2017; 11: 323-331Crossref PubMed Scopus (4) Google Scholar). Hence, yeast prions are low in complexity by definition, and prion-like domains are disordered FUS as a prion-like domain in by a search for proteins with sequence biases to yeast a of other proteins that are in SGs hnRNPA2, and M. Han T.W. Xie S. Shi K. Du X. Wu L.C. Mirzaei H. Goldsmith E.J. Longgood J. Pei J. Grishin N.V. Frantz D.E. Schneider J.W. Chen S. Li L. et al.Cell-free formation of RNA granules: Low complexity sequence domains form dynamic fibers within hydrogels.Cell. 2012; 149: 753-767Abstract Full Text Full Text PDF PubMed Scopus (1183) Google Scholar, S.M. Paul K.R. Ross E.D. Manipulating the aggregation activity of human prion-like proteins.Prion. 2017; 11: 323-331Crossref PubMed Scopus (4) Google Scholar, A. J. J. M. Kim Mittag T. Phase separation by low complexity domains stress assembly and Full Text Full Text PDF PubMed Scopus Google Scholar, Rosen M.K. and of by Full Text Full Text PDF PubMed Scopus Google Scholar). not LCDs are prion-like LCDs are common in proteins and to other LCDs J.C. X. RNA assembly of FUS Full Text Full Text PDF PubMed Scopus Google Scholar). the LCDs from hnRNPA2, FUS, and are phase transition and form hydrogels of amyloid-like M. Han T.W. Xie S. Shi K. Du X. Wu L.C. Mirzaei H. Goldsmith E.J. Longgood J. Pei J. Grishin N.V. Frantz D.E. Schneider J.W. Chen S. Li L. et al.Cell-free formation of RNA granules: Low complexity sequence domains form dynamic fibers within hydrogels.Cell. 2012; 149: 753-767Abstract Full Text Full Text PDF PubMed Scopus (1183) Google Scholar, A. L. S. M. S. J. S. A. M. et phase transition of the protein FUS by Full Text Full Text PDF PubMed Scopus Google Scholar, S. T. A. S. A. D. K. T. D. L. et domains in RNA proteins are for PubMed Scopus Google Scholar, T. S. A. D. Li et phase transition of FUS and reversible hydrogels hydrogels Full Text Full Text PDF PubMed Scopus Google Scholar). Phase transitions are also the and proteins contribute to MLO organization through their IDRs and LCDs B. L. Rosen M.K. disordered regions contribute interactions to 2018; Full Text Full Text PDF PubMed Scopus Google Scholar). The phase transition is a phase separation that to a phase with the proteins may a phase transition from to to form A. L. S. M. S. J. S. A. M. et phase transition of the protein FUS by Full Text Full Text PDF PubMed Scopus Google Scholar, T. S. A. D. Li et phase transition of FUS and reversible hydrogels hydrogels Full Text Full Text PDF PubMed Scopus Google Scholar, E.D. B. J. D.S. et and RNA form amyloid-like in 2018; PubMed Scopus Google Scholar, L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar). hnRNPA2, FUS, and have been found in and MLOs have been as a for formation in King J. granules as of PubMed Scopus Google Scholar, J. The of the proteins with prion-like domains in 2012; PubMed Scopus Google Scholar, phase in cell and 2017; PubMed Scopus Google Scholar). have been with and but are abundant of Examples include by Escherichia coli to prions in yeast to and protein granules in to D. 2019; PubMed Scopus Google Scholar, in 2012; PubMed Scopus Google Scholar). In these the of the of proteins to form on The from the LCDs of FUS, and are because these amyloid-like are M. Han T.W. Xie S. Shi K. Du X. Wu L.C. Mirzaei H. Goldsmith E.J. Longgood J. Pei J. Grishin N.V. Frantz D.E. Schneider J.W. Chen S. Li L. et al.Cell-free formation of RNA granules: Low complexity sequence domains form dynamic fibers within hydrogels.Cell. 2012; 149: 753-767Abstract Full Text Full Text PDF PubMed Scopus (1183) Google Scholar, S. T. A. S. A. D. K. T. D. L. et domains in RNA proteins are for PubMed Scopus Google Scholar, T. S. A. D. Li et phase transition of FUS and reversible hydrogels hydrogels Full Text Full Text PDF PubMed Scopus Google and by of or enables by LCDs to participate in the organization and of MLOs but with the of In we motifs termed low-complexity amyloid-like reversible kinked segments (LARKS) that the reversible of these proteins L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar). LARKS allow proteins to form structures that the to amyloid-like but the LARKS kinks in the which the with the we in that form and with their S. M. D. structures of PubMed Scopus Google Scholar). The LARKS is to form a but the of a and to function in the organization of MLOs. We searched for LARKS motifs in the human by protein sequences known LARKS structures and using a to a LARKS L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar, L. D. the proteins of amyloid-like S. A. PubMed Scopus Google Scholar). search through the human found that LARKS are common in LCDs of proteins found in MLOs. Here, we this search for LARKS in the organisms Mycobacterium tuberculosis, Saccharomyces cerevisiae, Plasmodium falciparum, and Drosophila to with the distribution of LARKS in the H. sapiens These model organisms because are and an of coli and M. are but M. is an P. as a S. cerevisiae as a and D. as an example of We the LARKS for these model organisms to that not have LARKS-rich LCD as the LCDs of S. cerevisiae and P. are not in with a of glycine in their LCD amino We on to of LARKS and amino structure. this to the that LARKS in We LARKS with LARKSdb found that the proteins with the most LARKS with proteins in L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar). We that LARKS are an LCDs of organisms that them to organize as in H. sapiens, and searched to LARKS-rich We defined a LARKS-rich protein as protein LARKS the for the of that species. We found that but not proteomes are rich in In H. sapiens, we that of the proteins are LARKS of proteins LCDs LARKS rich and from that proteins in H. sapiens are to be LARKS rich the proteins not have an we LARKS in LCD proteins from H. sapiens to other coli as a model with a that has few In fact, of coli proteins have an but that is in LARKS and to we D. and it has LCD and LARKS to H. D. proteins are also in LARKS we the S. cerevisiae which is a known for LCDs that form of the S. cerevisiae has and to are not in LARKS LARKS are a structural abundant in proteins of H. sapiens and D. but not in S. we other organisms with LCD and the The M. for and the P. for M. has a where of proteins have an and LARKS-rich proteins are in the proteins The found with P. where of proteins contain LCDs and of LARKS-rich proteins are to the LARKS proteomes of M. tuberculosis, D. melanogaster, and H. sapiens, we S. cerevisiae and P. to have LARKS In the that proteomes in their of LARKS LCD The aforementioned found the to which LARKS motifs are in proteins that contain an we the to which the LCDs of proteomes are in We also the to which proteins contain LCDs where LARKS do not in the do we the of and the of that are in an LCD and a LARKS and these LARKS LCD The of LARKS LCD is by the of to the of LARKS LCD in a We methods to a on the of the for the of LARKS LCD We this to the of this by the of LCD by the of LARKS the of LARKS LCD is the for the we the LCDs to be in of of of LCD LARKS LARKS LCD of in that found to be in or globular regions is in to of the The the of to the to the in the are the of the of the form of a with the of the the of the from of for The is the of LARKS by the of LCD the is the for the LARKS are in the is the LARKS are in LCDs with be for that The is to LARKS and LCD the LARKS LCD with the that LARKS are in LCDs in organisms P. falciparum. in a new The of in that found to be in or globular regions is in to of the The the of to the to the in the are the of the of the form of a with the of the the of the from of for The is the of LARKS by the of LCD the is the for the LARKS are in the is the LARKS are in LCDs with be for that The is to LARKS and LCD the LARKS LCD with the that LARKS are in LCDs in organisms P. falciparum. these results that for M. tuberculosis, D. melanogaster, and H. sapiens, the of that between LARKS and LCDs is by with the that LARKS are in LCDs and that the LARKS within the LCD in these S. cerevisiae a and P. has of LARKS in as the is within the with the from and by that LARKS in the LCDs of organisms that have LARKS LCD We searched for LARKS in LCDs because LCDs are and LARKS to be to allow interactions that LCDs are a reasonable proxy for are not always (5Wootton J.C. Non-globular domains in protein sequences: Automated segmentation using complexity measures.Comput. Chem. 1994; 18: 269-285Crossref PubMed Scopus (376) Google Scholar). we also the but for globular and disordered regions as defined by the protein sequences for and PubMed Scopus Google Scholar). We LARKS in globular regions LARKS and for the of LARKS is in globular regions of the LARKS motifs in IDRs of In fact, in proteomes that LARKS LCD we that LARKS are We this to LARKS are from globular regions the to be in IDRs that IDR is an LCD or gain LCDs in proteomes are in are we LCD amino of gain understanding of the proteomes of the of LARKS in we the amino of LCDs in We found glycine to be the most common in of glycine is with LARKS structures where the form kinks in the LARKS the L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar). In the proteomes with LARKS-rich LCDs M. tuberculosis, D. melanogaster, and H. we that glycine is the most common is not one of the most common in these proteomes but is the of the S. cerevisiae and P. falciparum. The of glycine in the amino acids of LARKS proteomes to absence in LARKS proteomes is gain the function of LARKS in we structures from LARKS-rich and proteins LARKS are structural motifs that interactions that form reversible and are with phase Here, we to that LARKS are motifs in LCDs and IDRs in H. we this to other we found a of LARKS in the LCDs of various species. proteomes LARKS are in globular regions that the to LARKS is in globular structure. LARKS are not in the LCDs of proteomes and and we the of LARKS-rich proteins with biology of the as by a of the in the LARKS-rich proteins of other proteomes we D. is most to H. sapiens in LARKS and LCD and to H. sapiens in are for LARKS LCD proteins between the proteomes are for proteins in RNA MLOs (e.g., and and formation and In of LARKS in the human we that are in LARKS and that phase separation may be an of formation L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar), and this in J. L. phase separation PubMed Scopus Google Scholar). for LARKS LCD proteins in D. a for of the which is the is a of the D. and in human be to LARKS to H. sapiens in structure. are from a and a of the the LARKS and sequences are the sequence and the in the protein have a are three structures of LARKS-rich from FUS, hnRNPA2, and and structures from and is with and the by protein with from the of a with from Drosophila with formation in and from the structures of proteins with dynamic MLOs hnRNPA2, and have LARKS and have kinks in their with the or low-complexity amyloid-like reversible kinked low-complexity in protein structure. example of a LARKS the kinked and between of and a of the kinked an example of a to the between the and are and because the from the structure. and of is a from a of LARKS or the proteins in as as structures in The LARKS-rich LCD and have found in and LARKS protein by low-complexity amyloid-like reversible kinked low-complexity S. cerevisiae not for LARKS LCD proteins in the but is to proteins with the most LARKS LCD are for mRNA P and SGs and to with that LARKS-rich proteins do not with proteins in S. cerevisiae We because are not LARKS LCD in S. cerevisiae, for proteins with LARKS LCD proteins with and may an of function of LCDs are not LARKS that is the the S. cerevisiae not for LARKS in but that proteins functions for phase separation as in H. sapiens but using motifs SGs are dynamic and human SGs S. S. D. L. D. S. interactions and protein the of PubMed Scopus Google Scholar), which from the LCDs in S. cerevisiae to motifs We the to do a of and LARKS in LCDs of H. sapiens and S. cerevisiae, but we a of proteins from to of LARKS and in these organisms and We the LARKS and of human prion-like proteins with LCDs hnRNPA2, TDP43, and FUS to the LCDs of yeast prion and The LCDs of the H. sapiens and the S. cerevisiae proteins have LARKS the protein as a and the of LARKS to we the of in the In H. sapiens prion-like the of is a yeast prion proteins LCDs with a a for The human prion-like proteins are also in LARKS with other or globular proteins which that of the of LARKS in human LCD proteins with yeast These from a and S. cerevisiae may to form but a is LCDs common on proteins in the LARKS may of The and in that most resembles S. cerevisiae is P. falciparum, which is a with LCDs that are in glycine and rich in asparagine for LARKS-rich proteins a for but we this may be and a few LCDs with a low of LARKS within them because P. has proteins with LARKS LCD The of M. has a of proteins that are with LARKS LCD and These proteins to the a of proteins with LCDs that the of the cell The of and 2017; PubMed Scopus Google Scholar). In fact, are with and cell A. T. H. M. D. In of 2018; Full Text Full Text PDF PubMed Scopus Google Scholar, M. bodies by and are with proteomes but in proteins that are for and in protein PubMed Scopus (4) Google Scholar). We it that the LARKS LCD proteins on an are to the is to that these LARKS-rich proteins may have with H. sapiens in to with LARKS-rich protein cells to help P. is also an but with M. tuberculosis, the is of LARKS in In the P. the is by a cell. in the the the within to it but is not to the with the M. which to with the cytoplasm The and of the Mycobacterium 18: PubMed Scopus Google Scholar). LARKS-rich proteins in are in and the LARKS-rich proteins on the cell be to with the have LCDs that in but as their and LARKS is their of is as the LARKS of LCDs may the and of that the LARKS-rich LCDs of human proteins to be like the yeast proteins S. S. D. L. D. S. interactions and protein the of PubMed Scopus Google Scholar), which like and are to form and We that this in be in LARKS and to this we of the of structures by and solid-state to the that LARKS have on structure. We to the structures of of proteins FUS and that have and with the of from that has LARKS The is a that to an that is the of found in The FUS and structures have a kinked that the formation of the of LARKS in a is be by the between and The in have a with LARKS where the kinks this to to The in FUS and structures are and in with their LARKS and The by LARKS are with amyloid-like L. D. L. T. D.S. structures of low-complexity protein segments kinked that assemble 2018; PubMed Scopus Google Scholar), the of FUS and M. Han T.W. Xie S. Shi K. Du X. Wu L.C. Mirzaei H. Goldsmith E.J. Longgood J. Pei J. Grishin N.V. Frantz D.E. Schneider J.W. Chen S. Li L. et al.Cell-free formation of RNA granules: Low complexity sequence domains form dynamic fibers within hydrogels.Cell. 2012; 149: 753-767Abstract Full Text Full Text PDF PubMed Scopus (1183) Google Scholar, T. S. A. D. Li et phase transition of FUS and reversible hydrogels hydrogels Full Text Full Text PDF PubMed Scopus Google Scholar, J. D. D.S. of the low-complexity domain of and to 11: PubMed Scopus Google Scholar). structural of and reversible and FUS (3Kato M. McKnight S.L. A solid-state conceptualization of information transfer from gene to message to protein.Annu. Rev. Biochem. 2018; 87: 351-390Crossref PubMed Scopus (62) Google to that LARKS in a that is with the biology of the are as is the case with from D. a to in is to the time K. of a in in PubMed Scopus Google Scholar). The sequence of an LCD that glutamine but has LARKS and The of a of glutamine a is by and this is in the of LARKS in the of LARKS with kinked structures within the are structures of segments of the FUS LCD and four of the LCD M. K.R. McKnight S.L. of FUS protein and to and phase separation of low-complexity 2017; Full Text Full Text PDF PubMed Scopus Google Scholar, M. K.R. M. and interactions within amyloid-like by a low-complexity protein sequence from 11: PubMed Scopus Google Scholar, P. D.S. structures of four 2019; PubMed Scopus Google Scholar). The segments with and kinked with as by and FUS and other proteins form hydrogels that be T. S. A. D. Li et phase transition of FUS and reversible hydrogels hydrogels Full Text Full Text PDF PubMed Scopus Google Scholar, J. D. D.S. of the low-complexity domain of and to 11: PubMed Scopus Google Scholar). may be that the kinked by LARKS may the form that may be a that to the In support the of LARKS as structural motifs in LCDs that help phase transitions and MLO organization in We LARKS abundant in that have dynamic MLOs (e.g., D. melanogaster, H. in that have dynamic SGs and abundant in proteins that are to the cytoplasm LARKS are not in LCDs (e.g., S. cerevisiae, P. and not in proteins that phase LARKS are one that organisms have for but not proteins that phase LARKS may the of the are in by interactions for phase separation aggregation by The to which structures form in MLOs Mittag T. in biomolecular PubMed Scopus Google Scholar), but we that amyloid-like structures rich in LARKS have kinked and the kinked structures the of protein by aggregation in MLOs proteins to S. A. PubMed Scopus Google Scholar). LARKS to be a structural through by species. the in this are permits to their proteins for LARKS proteomes used The proteomes and of 2017; M. S. P. D. and H. protein sequence with a not in the amino acids LARKS using the methods by et The computational methods identify segments that are to form LARKS the sequence of a new protein is six segments with an The for the sequence of the are a of a known LARKS and using a is is for three LARKS structures and and the segments is a for of the three the is to be a LARKS the of found in we within the LARKS as a LARKS in LCDs by using the (5Wootton J.C. Non-globular domains in protein sequences: Automated segmentation using complexity measures.Comput. Chem. 1994; 18: 269-285Crossref PubMed Scopus (376) Google Scholar). in a low in complexity as by it to be an LCD within a of low-complexity not to be in LCDs in IDRs using protein sequences for and PubMed Scopus Google Scholar). The and used to search proteomes for globular to be in a globular region using the to be and other to be in IDRs in to a or in and in be LARKS in LARKS in and LCD in of of (e.g., LARKS in by the of to be LARKS and in LCDs within a and this to an The found by the of in a (e.g., LARKS by the of in the We used to a for the to the for LARKS in we of where to the of LCD with and the of LARKS LCD from this to the LARKS LCD the of a distribution of the of LARKS LCD from the LCD The and to the in These the of the The found for by the of in one (e.g., by the of a in (e.g., to the LARKS The of this is a reasonable because are in the of the is or the for the we that protein to be or in LARKS in is to for the LARKS in LCD LARKS in and LCD in we the of LARKS the protein that has the in the for the LARKS of LARKS in a new The of proteins with an LCD and LARKS rich for to the by with from the to the of proteins with LCDs that from that that the of LARKS LCD proteins to a distribution of of LARKS LCD proteins that are found in the of the in to the of LCD The of LARKS LCD proteins and on the to it or within the are in the A by the of that that the of LARKS LCD proteins and by the of for P. where the of with LARKS LCD proteins to of LARKS LCD in LCD of LARKS LCD of LARKS LCD in a new on the of LARKS-rich proteins in with LARKS-rich defined as the of LARKS of a The of for these of proteins to the to A. H. B. K. A. A. A of protein and by PubMed Scopus Google Scholar). for and the of is in the are on the LARKSdb D. S. is and in other that have of with the of this We J. P. for support and and the of and and for The is the of the and not the of the of

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