Chemical tools to explore nutrient-driven O-GlcNAc cycling

Abstract

Posttranslational modifications (PTM) including glycosylation, phosphorylation, acetylation, methylation and ubiquitination dynamically alter the proteome. The evolutionarily conserved enzymes O-linked N-acetylglucosamine (O-GlcNAc) transferase (OGT) and O-GlcNAcase are responsible for the addition and removal, respectively, of the nutrient-sensitive PTM of protein serine and threonine residues with O-GlcNAc. Indeed, the O-GlcNAc modification acts at every step in the “central dogma” of molecular biology and alters signaling pathways leading to amplified or blunted biological responses. The cellular roles of OGT and the dynamic PTM O-GlcNAc have been clarified with recently developed chemical tools including high-throughput assays, structural and mechanistic studies and potent enzyme inhibitors. These evolving chemical tools complement genetic and biochemical approaches for exposing the underlying biological information conferred by O-GlcNAc cycling.     

Narrow endemic species Bellevalia webbiana shows significant intraspecific variation in tertiary CSR strategy

Insufficient attention has been devoted to inter-population and inter-individual variability of CSR strategy. Hence, we selected Bellevalia webbiana as a case study for answering the following questions: (1) is there a significant intraspecific variability in leaf traits determining a significant intraspecific variability in CSR parameters, even in a narrow endemic species with a relatively homogeneous habitat? (2) If yes, does this provide a significant intraspecific CSR strategy variation? For five populations, we calculated C, S, and R parameters using leaf area, leaf fresh weight, and leaf dry weight for 10 individuals each. We found that Bellevalia webbiana is a “CS” species considering single individuals, populations or the “whole species”. However, our data suggest a significant loss of information if the species is plotted as a single average point, and attest for population-based plasticity, highlighting the applicability of CSR theory at micro-evolutionary scale. When heterogeneity in coefficients of variation (CVs) is higher for the “whole species”, “deviant” populations should be retained as additional points. Hence, we advise to perform CSR analyses on more populations and verify the applicability of a single ternary set to represent the “whole species” using PERMANOVA, complemented by simple CV comparison for each parameter.     

Chemical approaches for investigating site-specific protein S-fatty acylation

Protein S-fatty acylation or S-palmitoylation is a reversible and regulated lipid post-translational modification (PTM) in eukaryotes. Loss-of-function mutagenesis studies have suggested important roles for protein S-fatty acylation in many fundamental biological pathways in development, neurobiology, and immunity that are also associated with human diseases. However, the hydrophobicity and reversibility of this PTM have made site-specific gain-of-function studies more challenging to investigate. In this review, we summarize recent chemical biology approaches and methods that have enabled site-specific gain-of-function studies of protein S-fatty acylation and the investigation of the mechanisms and significance of this PTM in eukaryotic biology.     

Blessed are the meek for they shall inherit the earth: quietness component of introversion is associated with single nucleotide polymorphisms in two circadian clock genes

Individual differences studied by chronobiologists and personality psychologists are usually shaped by polygenic selection occurring by small allele frequency shifts spreading across many loci. Therefore, the candidate gene association studies suffer from increased likelihood of false positive findings. We previously associated a PER3 single nucleotide polymorphism (SNP, rs228697) with self-ratings on personality-relevant nouns exemplifying personality dimension of Extraversion/Introversion in a sample of 88 female students. To replicate and extend this finding, we genotyped three more SNPs in three circadian clock genes. The results indicated that the minor alleles of PER3 rs228697 and PER2 rs934945 were rather similar in terms of their association with a personality type nicknamed “demure persona” (i.e. described by such nouns as “quietness”, “restraint”, “taciturnity”, “bashfulness”, “timidity”, “constraint”, and “reticence”). Analysis of data from populations of the 1000 Genomes Project suggested that, like frequencies of the minor alleles of many SNPs in circadian clock genes, the frequencies of these two SNPs were higher in populations of out-of-African ancestry compared to populations of African ancestry. We suggested that genetic candidates for Extraversion/Introversion can be prioritized in future association studies by means of identification of genetic signatures of polygenic selection imposed by out-of-Africa expansion of ancestral populations.     

Assessment of bacterial endosymbiont diversity in Otiorhynchusspp. (Coleoptera: Curculionidae) larvae using a multitag 454 pyrosequencing approach

Background

Weevils of the genus Otiorhynchus are regarded as devastating pests in a wide variety of horticultural crops worldwide. So far, little is known on the presence of endosymbionts in Otiorhynchus spp.. Investigation of endosymbiosis in this genus may help to understand the evolution of different reproductive strategies in these weevils (parthenogenesis or sexual reproduction), host-symbiont interactions, and may provide a future basis for novel pest management strategy development. Here, we used a multitag 454 pyrosequencing approach to assess the bacterial endosymbiont diversity in larvae of four economically important Otiorhynchus species.

Results

High-throughput tag-encoded FLX amplicon pyrosequencing of a bacterial 16S rDNA fragment was used to characterise bacterial communities associated with different Otiorhynchus spp. larvae. By sequencing a total of ~48,000 PCR amplicons, we identified 49 different operational taxonomic units (OTUs) as bacterial endosymbionts in the four studied Otiorhynchus species. More than 90% of all sequence reads belonged either to the genus Rickettsia or showed homology to the phylogenetic group of “Candidatus Blochmannia” and to endosymbionts of the lice Pedicinus obtusus and P. badii. By using specific primers for the genera Rickettsia and “Candidatus Blochmannia”, we identified a new phylogenetic clade of Rickettsia as well as “Candidatus Nardonella” endosymbionts in Otiorhynchus spp. which are closely related to “Candidatus Blochmannia” bacteria.

Conclusions

Here, we used multitag 454 pyrosequencing for assessment of insect endosymbiotic communities in weevils. As 454 pyrosequencing generates only quite short sequences, results of such studies can be regarded as a first step towards identifying respective endosymbiotic species in insects. In the second step of our study, we analysed sequences of specific gene regions for a more detailed phylogeny of selected endosymbiont genera. As a result we identified the presence of Rickettsia and “Candidatus Nardonella” endosymbionts in Otiorhynchus spp.. This knowledge is an important step in exploring bacteria-insect associations for potential use in insect pest control.

     

Simultaneous Quantification of the Acetylome and Succinylome by ‘One‐Pot’ Affinity Enrichment

Protein posttranslational modifications (PTMs) are of increasing interest in biomedical research, yet studies rarely examine more than one PTM. One barrier to multi‐PTM studies is the time cost for both sample preparation and data acquisition, which scale linearly with the number of modifications. The most prohibitive requirement is often the need for large amounts of sample, which must be increased proportionally with the number of PTM enrichment steps. Here, a streamlined, quantitative label‐free proteomic workflow—“one‐pot” PTM enrichment—that enables comprehensive identification and quantification of peptides containing acetylated and succinylated lysine residues from a single sample containing as little as 1 mg mitochondria protein is described. Coupled with a label‐free, data‐independent acquisition (DIA), 2235 acetylated and 2173 succinylated peptides with the one‐pot method are identified and quantified and peak areas are shown to be highly correlated between the one‐pot and traditional single‐PTM enrichments. The ‘one‐pot’ method makes possible detection of multiple PTMs occurring on the same peptide, and it is shown that it can be used to make unique biological insights into PTM crosstalk. Compared to single‐PTM enrichments, the one‐pot workflow has equivalent reproducibility and enables direct assessment of PTM crosstalk from biological samples in less time from less tissue.     

Compatible solute addition to biological systems treating waste/wastewater to counteract osmotic and other environmental stresses: a review

This study reviews the addition of compatible solutes to biological systems as a strategy to counteract osmolarity and other environmental stresses. At high osmolarity many microorganisms accumulate organic solutes called “compatible solutes” in order to balance osmotic pressure between the cytoplasm and the environment. These organic compounds are called compatible solutes because they can function inside the cell without the need for special adaptation of the intracellular enzymes, and also serve as protein stabilizers in the presence of high ionic strength. Moreover, the compatible solutes strategy is regularly being employed by the cell, not only under osmotic stress at high salinity, but also under other extreme environmental conditions such as low temperature, freezing, heat, starvation, dryness, recalcitrant compounds and solvent stresses. The accumulation of these solutes from the environment has energetically a lower cost than de novo synthesis. Based on this cell mechanism several studies in the field of environmental biotechnology (most of them on biological wastewater treatment) employed this strategy by exogenously adding compatible solutes to the wastewater or medium in order to alleviate environmental stress. This current paper critically reviews and evaluates these studies, and examines the future potential of this approach. In addition to this, a strategy for the successful implementation of compatible solutes in biological systems is proposed.     

Patterns on a parr: Drivers of long‐term salmon parr length in U.K. and French rivers depend on geographical scale

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The promise of primatology fulfilled?

In 1972, Sherwood Washburn, one of the forerunners of biological anthropology, gave an invited address during the 4th Congress of the International Primatological Society in Portland, Oregon, in which he expounded his vision for the field of primatology. His address was published the following year in the American Journal of Physical Anthropology and titled: “The promise of primatology.” In this centennial commentary, we revisit Washburn's “promise”, 45 years on. His address and article discuss the constraints acting on the field, including a positioning of the discipline across different kinds of university departments, and within the social sciences, which he viewed as a mixed blessing. Prescient aspects of Washburn's address include a focus on the need to study communication multimodally, and a hope that the study of mechanisms would become foundational within the field. We discuss new promising aspects of primatology, focusing on technological advances in a number of areas highlighted by Washburn that have ushered in new eras of research, and the increasingly large number of long‐term field sites, which see the discipline well‐set for new developmental and longitudinal studies. We find much to admire in Washburn's keen foresight, and natural intuition. Washburn hoped that primatology would repudiate the notion that “the social should be studied without reference to the biological.” In this regard, we consider much of Washburn's promise fulfilled.     

Investigation and identification of functional post-translational modification sites associated with drug binding and protein-protein interactions

Background

Protein post-translational modification (PTM) plays an essential role in various cellular processes that modulates the physical and chemical properties, folding, conformation, stability and activity of proteins, thereby modifying the functions of proteins. The improved throughput of mass spectrometry (MS) or MS/MS technology has not only brought about a surge in proteome-scale studies, but also contributed to a fruitful list of identified PTMs. However, with the increase in the number of identified PTMs, perhaps the more crucial question is what kind of biological mechanisms these PTMs are involved in. This is particularly important in light of the fact that most protein-based pharmaceuticals deliver their therapeutic effects through some form of PTM. Yet, our understanding is still limited with respect to the local effects and frequency of PTM sites near pharmaceutical binding sites and the interfaces of protein-protein interaction (PPI). Understanding PTM’s function is critical to our ability to manipulate the biological mechanisms of protein.

Results

In this study, to understand the regulation of protein functions by PTMs, we mapped 25,835 PTM sites to proteins with available three-dimensional (3D) structural information in the Protein Data Bank (PDB), including 1785 modified PTM sites on the 3D structure. Based on the acquired structural PTM sites, we proposed to use five properties for the structural characterization of PTM substrate sites: the spatial composition of amino acids, residues and side-chain orientations surrounding the PTM substrate sites, as well as the secondary structure, division of acidity and alkaline residues, and solvent-accessible surface area. We further mapped the structural PTM sites to the structures of drug binding and PPI sites, identifying a total of 1917 PTM sites that may affect PPI and 3951 PTM sites associated with drug-target binding. An integrated analytical platform (CruxPTM), with a variety of methods and online molecular docking tools for exploring the structural characteristics of PTMs, is presented. In addition, all tertiary structures of PTM sites on proteins can be visualized using the JSmol program.

Conclusion

Resolving the function of PTM sites is important for understanding the role that proteins play in biological mechanisms. Our work attempted to delineate the structural correlation between PTM sites and PPI or drug-target binding. CurxPTM could help scientists narrow the scope of their PTM research and enhance the efficiency of PTM identification in the face of big proteome data. CruxPTM is now available at http://csb.cse.yzu.edu.tw/CruxPTM/.

     

Molecular basis for Poria cocos mushroom polysaccharide used as an antitumour drug in China

Poria cocos is an edible medicinal fungus known as “Fuling” in Chinese and has been used as a Chinese traditional medicine for more than two thousand years. Pharmacological studies reveal that polysaccharide is the most abundant substance in Poria cocos and has a wide range of biological activities including antitumour, immunomodulation, anti‐inflammation, antioxidation, anti‐ageing, antihepatitis, antidiabetics and anti‐haemorrhagic fever effects. As a result, “Poria cocos polysaccharide oral solution” was developed and sold as an over‐the‐counter health supplement since 1970s. In 2015, “Polysaccharidum of Poria cocos oral solution” was approved as a drug by Chinese Food and Drug Administration for treating multiple types of cancers, hepatitis and other diseases alone or during chemo‐ or radiation therapy for patients with cancer. In this article, biochemical, preclinical and clinical studies of Poria cocos polysaccharide from 72 independent studies during the past 46 years (1970‐2016) based on PubMed, VIP (Chongqing VIP Chinese Scientific Journals Database), CNKI (China National Knowledge Infrastructure) and Wanfang database searches are summarized. The structure, pharmacological effects, clinical efficacy, immunobalancing molecular mechanism and toxicity of Poria cocos polysaccharide are deliberated to provide a general picture of Poria cocos polysaccharide as a clinically used antitumour drug.     

Reproductive isolation due to prezygotic isolation and postzygotic cytoplasmic incompatibility in parasitoid wasps

The reproductive barriers that prevent gene flow between closely related species are a major topic in evolutionary research. Insect clades with parasitoid lifestyle are among the most species‐rich insects and new species are constantly described, indicating that speciation occurs frequently in this group. However, there are only very few studies on speciation in parasitoids. We studied reproductive barriers in two lineages of Lariophagus distinguendus (Chalcidoidea: Hymenoptera), a parasitoid wasp of pest beetle larvae that occur in human environments. One of the two lineages occurs in households preferably attacking larvae of the drugstore beetle Stegobium paniceum (“DB‐lineage”), the other in grain stores with larvae of the granary weevil Sitophilus granarius as main host (“GW‐lineage”). Between two populations of the DB‐lineage, we identified slight sexual isolation as intraspecific barrier. Between populations from both lineages, we found almost complete sexual isolation caused by female mate choice, and postzygotic isolation, which is partially caused by cytoplasmic incompatibility induced by so far undescribed endosymbionts which are not Wolbachia or Cardinium. Because separation between the two lineages is almost complete, they should be considered as separate species according to the biological species concept. This demonstrates that cryptic species within parasitoid Hymenoptera also occur in Central Europe in close contact to humans.     

The manipulation mechanism of “push–pull” habitat management strategy and advances in its application

The “push–pull” habitat management strategy, as a new powerful and effective tool in integrated pest management (IPM), uses a combination of behavior-modifying stimuli to manipulate the distribution and abundance of pests and/or their natural enemies for pest control. In the “push–pull” strategy, pests are repelled or deterred away from the protected resource (push) by stimuli that disturb the host location and modulate the host become unattractive or unsuitable for the feeding and oviposition of pests. By using highly attractive stimuli, the target pests are simultaneously attracted (pull) to the specific source in which they are subsequently concentrated, facilitating their elimination and leaving the protected resource. Since stimuli usually are nontoxic, either “push” or “pull” components, the strategy is usually companied by population-reduced methods, such as using insecticides, exploiting nature enemies, and placing traps. Among them, methods of biological control are the most preferred. The “push–pull” strategies maximize efficacy of behavior-modifying stimuli through the additive and synergistic effects of integrating the use of methods for population reduction. In this review, the principles of the “push–pull” strategy were firstly described, then the potential behavior-modifying stimuli for “push” and “pull” components were introduced. The stimuli for use in “push–pull” strategies primarily include visual cues and chemical cues (synthetic or plant- or insect-derived semiochemicals). Visual stimuli, repellent and trap plants, host and non-host volatiles, insect pheromones, and antifeedants and oviposition deterrents are usually applied as the potential stimuli in the “push–pull” strategy for pest control. The stimuli are grouped as long-range stimuli and short-range stimuli. In addition, we also summarize models of “push–pull” habitat management strategy, such as agriculture, horticulture and forestry, of pest control and some successful case studies in applying of “push–pull” strategy and its potential ecological benefits. The “push–pull” technology (PPT), so far the most effective “push–pull” strategy in practice by farmers, developed for management of both stemborers and Striga hermonthica in maize-based farming system in eastern Africa were reviewed. The PPT uses an intercrop of repellent plants and border crops of attractive trap plants. Stemborer moths are effectively repelled away from the maize crop (push) by Desmodium and molasses grass, and are subsequently attracted (or trapped) to (pull) by the Napier grass and Sudan grass which emit the green leaf volatiles (GLVs) showing attractant properties. Finally, the problems in current research and future perspectives of the “push–pull” habitat management strategy are discussed in the present paper.     

Sequence polymorphisms in wild,weedy, and cultivated rice suggest seed‐shattering locus sh4 played a minor role in Asian rice domestication

The predominant view regarding Asian rice domestication is that the initial origin of nonshattering involved a single gene of large effect, specifically, the sh4 locus via the evolutionary replacement of a dominant allele for shattering with a recessive allele for reduced shattering. Data have accumulated to challenge this hypothesis. Specifically, a few studies have reported occasional seed‐shattering plants from populations of the wild progenitor of cultivated rice (Oryza rufipogon complex) being homozygous for the putative “nonshattering” sh4 alleles. We tested the sh4 hypothesis for the domestication of cultivated rice by obtaining genotypes and phenotypes for a diverse set of samples of wild, weedy, and cultivated rice accessions. The cultivars were fixed for the putative “nonshattering” allele and nonshattering phenotype, but wild rice accessions are highly polymorphic for the putative “nonshattering” allele (frequency ~26%) with shattering phenotype. All weedy rice accessions are the “nonshattering” genotype at the sh4 locus but with shattering phenotype. These data challenge the widely accepted hypothesis that a single nucleotide mutation (“G”/“T”) of the sh4 locus is the major driving force for rice domestication. Instead, we hypothesize that unidentified shattering loci are responsible for the initial domestication of cultivated rice through reduced seed shattering.     

Phosphoproteomic analysis of wild‐type and antimony‐resistant Leishmania braziliensis lines by 2D‐DIGE technology

Protein phosphorylation is one of the most studied post‐translational modifications that is involved in different cellular events in Leishmania. In this study, we performed a comparative phosphoproteomics analysis of potassium antimonyl tartrate (SbIII)‐resistant and ‐susceptible lines of Leishmania braziliensis using a 2D‐DIGE approach followed by MS. In order to investigate the differential phosphoprotein abundance associated with the drug‐induced stress response and SbIII‐resistance mechanisms, we compared nontreated and SbIII‐treated samples of each line. Pair wise comparisons revealed a total of 116 spots that showed a statistically significant difference in phosphoprotein abundance, including 11 and 34 spots specifically correlated with drug treatment and resistance, respectively. We identified 48 different proteins distributed into seven biological process categories. The category “protein folding/chaperones and stress response” is mainly implicated in response to SbIII treatment, while the categories “antioxidant/detoxification,” “metabolic process,” “RNA/DNA processing,” and “protein biosynthesis” are modulated in the case of antimony resistance. Multiple sequence alignments were performed to validate the conservation of phosphorylated residues in nine proteins identified here. Western blot assays were carried out to validate the quantitative phosphoproteome analysis. The results revealed differential expression level of three phosphoproteins in the lines analyzed. This novel study allowed us to profile the L. braziliensis phosphoproteome, identifying several potential candidates for biochemical or signaling networks associated with antimony resistance phenotype in this parasite.     

Disrupted montane forest recovery hinders biodiversity conservation in the tropical Andes

Aim

Andean montane forests are biodiversity hotspots and large carbon stores and they provide numerous ecosystem services. Following land abandonment after centuries of forest clearing for agriculture in the Andes, there is an opportunity for forest recovery. Field-based studies show that forests do not always recover. However, large-scale and long-term knowledge of recovery dynamics of Andean forests remains scarce. This paper analyses tropical montane forest recovery trajectories over a 15-year time frame at the landscape and tropical Andean scale to inform restoration planning.

Methods

We first detect “potential recovery” as areas that have experienced a forest transition between 2000 and 2005. Then, we use Landsat time series analysis of the normalized difference water index (NDWI) to classify four “realized recovery” trajectories (“ongoing”, “arrested”, “disrupted” and “no recovery”) based on a sequential pattern of 5-yearly Z-score anomalies for 2005–2020. We compare these results against an analysis of change in tree cover to validate against other datasets.

Results

Across the tropical Andes, we detected a potential recovery area of 274 km² over the period. Despite increases in tree cover, most areas of the Andes remained in early successional states (10–25% tree cover), and NDWI levelled out after 5–10 years. Of all potential forest recovery areas, 22% showed “ongoing recovery”, 61% showed either “disrupted” or “arrested recovery”, and 17% showed “no recovery”. Our method captured forest recovery dynamics in a Peruvian arrested succession context and in landscape-scale tree-planting efforts in Ecuador.

Main conclusions

Forest recovery across the Andes is mostly disrupted, arrested or unsuccessful, with consequences for biodiversity recovery and provision of ecosystem services. Low-recovery areas identified in this study might be good candidates for active restoration interventions in this UN Decade on Restoration. Future studies could determine restoration strategies and priorities and suggest management strategies at a local planning scale across key regions in the biodiversity hotspot.     

Phylogenetic relationships in the Sceloporus variabilis (Squamata: Phrynosomatidae) complex based on three molecular markers,continuous characters and geometric morphometric data

The monophyly of the Sceloporus variabilis group is well established with five species and two species complexes, but phylogenetic relationships within species complexes are still uncertain. We studied 278 specimens in 20 terminals to sample all taxa in the “variabilis group,” including three subspecies in the “variabilis complex,” and two outgroups (Sceloporus grammicus and Sceloporus megalepidurus). We assembled an extensive morphological data set with discrete and continuous characters (distances and scale counts), including geometric morphometric data (landmark coordinates of three shapes), and a three‐marker molecular data set as well (ND4, 12S and RAG1). We conducted parsimony and Bayesian phylogenetic inferences on these data, including several partitioning and weighting schemes. We suggest elevating three subspecies to full species status. Therefore, we recommend recognition of nine species in the “variabilis group.” First, S. variabilis is sister to Sceloporus teapensis. In turn, Sceloporus cozumelae is sister to Sceloporus olloporus. These four species are a monophyletic group, which is sister to Sceloporus smithi. Finally, Sceloporus marmoratus is sister of the clade of five species. The other species in the “variabilis group” (Sceloporus chrysostictus, Sceloporus couchii and Sceloporus parvus) are a paraphyletic grade at the base of the tree. Our analyses reject the existence of the “variabilis complex.” We conducted a parsimony‐based ancestral reconstruction on body size (snout–vent length), femoral pores and dorsal scales and related morphological changes to geographic distribution of the species. Our phylogenetic hypothesis will allow best designs of comparative studies with species in the “variabilis group,” one of the earliest divergent lineages in the genus.     

The rhetoric of Islamophobia: an analysis of the means of persuasion in Hege Storhaug’s writings on Islam and Muslims

Few actors have had a greater impact on the “framing of Muslims” as a social and political “problem” in Norway since 2001 than Hege Storhaug of the government- and corporate billionaire funded civil society organization Human Rights Service (HRS). Using the methodological tools of the “rhetorical branch” of Critical Discourse Analysis (CDA), and applying the Aristotelian concepts of ethos, logos and pathos, we analyze the bestselling popular title on Islam and Muslims ever published in Norway, namely Storhaug’s self-published 2015 title “Islam – The Eleventh Plague”. We argue that Storhaug’s popular success must be understood in light of her rhetorical appeals to femonationalism, the critique of religion and “Enlightenment” values. We show how she in her writings incites fear of the Muslim “Other” through specific rhetorical devices and a positioning of herself as a defender of the “nation” and the “people” – against national and international “elites”.     

Examining post‐translational modification‐mediated protein–protein interactions using a chemical proteomics approach

Post‐translational modifications (PTM) of proteins can control complex and dynamic cellular processes via regulating interactions between key proteins. To understand these regulatory mechanisms, it is critical that we can profile the PTM‐dependent protein–protein interactions. However, identifying these interactions can be very difficult using available approaches, as PTMs can be dynamic and often mediate relatively weak protein–protein interactions. We have recently developed CLASPI (cross‐linking‐assisted and stable isotope labeling in cell culture‐based protein identification), a chemical proteomics approach to examine protein–protein interactions mediated by methylation in human cell lysates. Here, we report three extensions of the CLASPI approach. First, we show that CLASPI can be used to analyze methylation‐dependent protein–protein interactions in lysates of fission yeast, a genetically tractable model organism. For these studies, we examined trimethylated histone H3 lysine‐9 (H3K9Me₃)‐dependent protein–protein interactions. Second, we demonstrate that CLASPI can be used to examine phosphorylation‐dependent protein–protein interactions. In particular, we profile proteins recognizing phosphorylated histone H3 threonine‐3 (H3T3‐Phos), a mitotic histone “mark” appearing exclusively during cell division. Our approach identified survivin, the only known H3T3‐Phos‐binding protein, as well as other proteins, such as MCAK and KIF2A, that are likely to be involved in weak but selective interactions with this histone phosphorylation “mark”. Finally, we demonstrate that the CLASPI approach can be used to study the interplay between histone H3T3‐Phos and trimethylation on the adjacent residue lysine 4 (H3K4Me₃). Together, our findings indicate the CLASPI approach can be broadly applied to profile protein–protein interactions mediated by PTMs.