Abstract
Decoding the role of histone posttranslational modifications (PTMs) is key to understand the fundamental process of epigenetic regulation. This is well studied for PTMs of core histones but ...not for linker histone H1 in general and its ubiquitylation in particular due to a lack of proper tools. Here, we report on the chemical synthesis of site-specifically mono-ubiquitylated H1.2 and identify its ubiquitin-dependent interactome on a proteome-wide scale. We show that site-specific ubiquitylation of H1 at position K64 modulates interactions with deubiquitylating enzymes and the deacetylase
SIRT1
. Moreover, it affects H1-dependent chromatosome assembly and phase separation resulting in a more open chromatosome conformation generally associated with a transcriptionally active chromatin state. In summary, we propose that site-specific ubiquitylation plays a general regulatory role for linker histone H1.
Abstract
The nucleotides diadenosine triphosphate (Ap
3
A) and diadenosine tetraphosphate (Ap
4
A) are formed in prokaryotic and eukaryotic cells. Since their concentrations increase significantly ...upon cellular stress, they are considered to be alarmones triggering stress adaptive processes. However, their cellular roles remain elusive. To elucidate the proteome-wide interactome of Ap
3
A and Ap
4
A and thereby gain insights into their cellular roles, we herein report the development of photoaffinity-labeling probes and their employment in chemical proteomics. We demonstrate that the identified Ap
n
A interactors are involved in many fundamental cellular processes including carboxylic acid and nucleotide metabolism, gene expression, various regulatory processes and cellular response mechanisms and only around half of them are known nucleotide interactors. Our results highlight common functions of these Ap
n
As across the domains of life, but also identify those that are different for Ap
3
A or Ap
4
A. This study provides a rich source for further functional studies of these nucleotides and depicts useful tools for characterization of their regulatory mechanisms in cells.
Oocyte maturation and early embryo development occur in vertebrates in the near absence of transcription. Thus, sexual reproduction of vertebrates critically depends on the timely translation of ...mRNAs already stockpiled in the oocyte. Yet how translational activation of specific mRNAs is temporally coordinated is still incompletely understood. Here, we elucidate the function of Zar1l, a yet uncharacterized member of the Zar RNA-binding protein family, in Xenopus oocytes. Employing TRIM-Away, we demonstrate that loss of Zar1l accelerates hormone-induced meiotic resumption of Xenopus oocytes due to premature accumulation of the M-phase-promoting kinase cMos. We show that Zar1l is a constituent of a large ribonucleoparticle containing the translation repressor 4E-T and the central polyadenylation regulator CPEB1, and that it binds directly to the cMos mRNA. Partial, hormone-induced degradation of Zar1l liberates 4E-T from CPEB1, which weakens translational repression of mRNAs encoding cMos and likely additional M-phase-promoting factors. Thus, our study provides fundamental insights into the mechanisms that ensure temporally regulated translation of key cell cycle regulators during oocyte maturation, which is essential for sexual reproductivity.
The abnormal amplification of a CAG repeat in the gene coding for huntingtin (HTT) leads to Huntington's disease (HD). At the protein level, this translates into the expansion of a polyglutamine ...(polyQ) stretch located at the HTT N terminus, which renders HTT aggregation prone by unknown mechanisms. Here we investigated the effects of polyQ expansion on HTT in a complex with its stabilizing interaction partner huntingtin-associated protein 40 (HAP40). Surprisingly, our comprehensive biophysical, crosslinking mass spectrometry and cryo-EM experiments revealed no major differences in the conformation of HTT-HAP40 complexes of various polyQ length, including 17QHTT-HAP40 (wild type), 46QHTT-HAP40 (typical polyQ length in HD patients), and 128QHTT-HAP40 (extreme polyQ length). Thus, HTT polyQ expansion does not alter the global conformation of HTT when associated with HAP40.
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•Expansion of HTT polyQ repeat leads to HTT aggregation in Huntington's disease•HTT is stabilized within the HTT-HAP40 complex, enabling structural studies•HTT polyQ expansion does not alter the stability of the HTT-HAP40 complex•The structure of the HTT-HAP40 complex is not influenced by polyQ length
The expansion of a polyglutamine (polyQ) repeat in huntingtin (HTT) causes HTT aggregation in Huntington's disease by unknown mechanisms. Huang et al. combine biophysical methods, cryo-EM, and crosslinking mass spectrometry to show that polyQ expansion does not alter HTT structure within the HTT-HAP40 complex.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Intracellular dinucleoside polyphosphates (NpnNs) have been known for decades but the functional role remains enigmatic. Diadenosine triphosphate (Ap3A) is one of the most prominent examples, and its ...intercellular concentration was shown to increase upon cellular stress. By employment of previously reported Ap3A‐based photoaffinity‐labeling probes (PALPs) in chemical proteomics, we investigated the Ap3A interactome in the human lung carcinoma cell line H1299. The cell line is deficient of the fragile histidine triade (Fhit) protein, a hydrolase of Ap3A and tumor suppressor. Overall, the number of identified potential interaction partners was significantly lower than in the previously investigated HEK293T cell line. Gene ontology term analysis revealed that the identified proteins participate in similar pathways as for HEK293T, but the percentage of proteins involved in RNA‐related processes is higher for H1299. The obtained results highlight similarities and differences of the Ap3A interaction network in different cell lines and give further indications regarding the importance of the presence of Fhit.
The protein interaction network of the alarmone diadenosine triphosphate (Ap3A) in the cancer cell line H1299 was determined by chemical proteomics. Diazirine photoaffinity probes were used to covalently label interacting proteins with the affinity tag desthiobiotin, and the proteins were identified by their respective peptides via affinity purification coupled to mass spectrometry.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
We calculate spin, charge, and azimuthal asymmetries in deeply virtual Compton scattering at leading twist-two level. The measurement of these asymmetries gives access to the imaginary and real part ...of all deeply virtual Compton scattering amplitudes. We note that a consistent description of this process requires taking into account twist-three contributions and we give then a model dependent estimate of these asymmetries.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
We study the amplitude of deeply virtual Compton scattering in next-to-leading order of perturbation theory including the two-loop evolution effects for different sets of skewed parton distributions ...(SPDs). It turns out that in the minimal subtraction scheme the relative radiative corrections are typically of order 20–50%. In special cases they can even be much larger. We analyze the dependence of our predictions on the choice of SPD, that will allow to discriminate between possible models of SPDs from future high precision experimental data, and discuss shortly theoretical uncertainties induced by radiative corrections.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
We present a general method for the solution of the renormalization group equations for the non-forward parton distributions on the two-loop level in the flavour singlet channel based on an ...orthogonal polynomial reconstruction. Using this formalism we study the effects of the evolution on recently proposed model distribution functions.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
We study the effects of next-to-leading order corrections on the evolution of the twist-two non-forward parton distribution functions in the flavour non-singlet sector. It is found that the deviation ...from leading order evolution is small for all values of the parton momentum fraction variable for moderately large values of the scale parameter.
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IJS, IMTLJ, KILJ, KISLJ, NUK, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
The techniques researched, developed and applied towards the measurement of radioisotope concentrations at ultra-low levels in the real-time solar neutrino experiment BOREXINO at Gran Sasso are ...presented and illustrated with specific results of widespread interest. We report the use of low-level germanium gamma spectrometry, low-level miniaturized gas proportional counters and low background scintillation detectors developed in solar neutrino research. Each now sets records in its field. We additionally describe our techniques of radiochemical ultra-pure, few atom manipulations and extractions. Forefront measurements also result from the powerful combination of neutron activation and low-level counting. Finally, with our techniques and commercially available mass spectrometry and atomic absorption spectroscopy, new low-level detection limits for isotopes of interest are obtained.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
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