Bio-molecular architects: a scaffold provided by the C-terminal domain of eukaryotic RNA polymerase II

In eukaryotic cells, the transcription of genes is accurately orchestrated both spatially and temporally by the C-terminal domain of RNA polymerase II (CTD). The CTD provides a dynamic platform to recruit different regulators of the transcription apparatus. Different posttranslational modifications are precisely applied to specific sites of the CTD to coordinate transcription process. Regulators of the RNA polymerase II must identify specific sites in the CTD for cellular survival, metabolism, and development. Even though the CTD is disordered in the eukaryotic RNA polymerase II crystal structures due to its intrinsic flexibility, recent advances in the complex structural analysis of the CTD with its binding partners provide essential clues for understanding how selectivity is achieved for individual site recognition. The recent discoveries of the interactions between the CTD and histone modification enzymes disclose an important role of the CTD in epigenetic control of the eukaryotic gene expression. The intersection of the CTD code with the histone code discloses an intriguing yet complicated network for eukaryotic transcriptional regulation

Efficient and robust preparation of tyrosine phosphorylated intrinsically disordered proteins

Intrinsically disordered proteins (IDPs) are subject to post-translational modifications. This allows the same polypeptide to undertake different interaction networks with different consequences, ranging from regulatory signalling networks to formation of membraneless organelles. We report a robust method for co-expression of modification enzyme and SUMO-tagged IDP with subsequent purification procedure allowing production of modified IDP. The robustness of our protocol is demonstrated on a challenging system, RNA polymerase II C-terminal domain (CM), that is a low-complexity repetitive region with multiple phosphorylation sites. In vitro phosphorylation approaches fail to yield multiple-site phosphorylated CTD, whereas our in vivo protocol allows to rapidly produce near homogeneous phosphorylated CTD at a low cost. These samples can be used in functional and structural studies

Ctk1 promotes dissociation of basal transcription factors from elongating RNA polymerase II

As RNA polymerase II (RNApII) transitions from initiation to elongation, Mediator and the basal transcription factors TFIID, TFIIA, TFIIH, and TFIIE remain at the promoter as part of a scaffold complex, whereas TFIIB and TFIIF dissociate. The yeast Ctk1 kinase associates with elongation complexes and phosphorylates serine 2 in the YSPTSPS repeats of the Rpb1 C-terminal domain, a modification that couples transcription to mRNA 3′-end processing. The higher eukaryotic kinase Cdk9 not only performs a similar function, but also functions at the 5′-end of genes in the transition from initiation to elongation. In strains lacking Ctk1, many basal transcription factors cross-link throughout transcribed regions, apparently remaining associated with RNApII until it terminates. Consistent with this observation, preinitiation complexes formed on immobilized templates with transcription extracts lacking Ctk1 leave lower levels of the scaffold complex behind after escape. Taken together, these results suggest that Ctk1 is necessary for the release of RNApII from basal transcription factors. Interestingly, this function of Ctk1 is independent of its kinase activity, suggesting a structural function of the protein

Thiol-Functionalized Mesoporous Silica for Effective Trap of Mercury in Rats

The chance of exposure to heavy metal for human being rises severely today due to the increasing water contamination and air pollution. Here, we prepared a series of thiol-functionalized mesoporous silica as oral formulation for the prevention and treatment of heavy metal poisoning. The successful incorporation of thiol was verified by the FTIR spectra. SBA15-SH-10 was used for the study as it is of uniform mesopores and fine water dispersibility. In simulated gastrointestinal fluid, the thiol-functionalized mesoporous silica can selectively capture heavy metal, showing a very high affinity for inorganic mercury (II). The blood and urine mercury levels of rats fed with a diet containing Hg (II) and material were significantly lower than those of rats fed with the metal-rich diet only. On the contrary, the mercury content in fecal excretion of the treatment group increased more than twice as much as that of the control group. This result indicated that SBA15-SH-10 could effectively remove mercury (II) in vivo and the mercury loaded on SBA15-SH-10 would be excreted out. Hence, SBA15-SH-10 has potential application in preventing and treating heavy metal poisoning via digestive system

Comparison of Short-Wavelength Reduced-Illuminance and Conventional Autofluorescence Imaging in Stargardt Macular Dystrophy

Purpose To compare grading results between short-wavelength reduced-illuminance and conventional autofluorescence imaging in Stargardt macular dystrophy. Design Reliability study. Methods setting: Moorfields Eye Hospital, London (United Kingdom). patients: Eighteen patients (18 eyes) with Stargardt macular dystrophy. observation procedures: A series of 3 fundus autofluorescence images using 3 different acquisition parameters on a custom-patched device were obtained: (1) 25% laser power and total sensitivity 87; (2) 25% laser power and freely adjusted sensitivity; and (3) 100% laser power and freely adjusted total sensitivity (conventional). The total area of 2 hypoautofluorescent lesion types (definitely decreased autofluorescence and poorly demarcated questionably decreased autofluorescence) was measured. main outcome measures: Agreement in grading between the 3 imaging methods was assessed by kappa coefficients (κ) and intraclass correlation coefficients. Results The mean ± standard deviation area for images acquired with 25% laser power and freely adjusted total sensitivity was 2.04 ± 1.87 mm2 for definitely decreased autofluorescence (n = 15) and 1.86 ± 2.14 mm2 for poorly demarcated questionably decreased autofluorescence (n = 12). The intraclass correlation coefficient (95% confidence interval) was 0.964 (0.929, 0.999) for definitely decreased autofluorescence and 0.268 (0.000, 0.730) for poorly demarcated questionably decreased autofluorescence. Conclusions Short-wavelength reduced-illuminance and conventional fundus autofluorescence imaging showed good concordance in assessing areas of definitely decreased autofluorescence. However, there was significantly higher variability between imaging modalities for assessing areas of poorly demarcated questionably decreased autofluorescence

Challenges, Technologies and Solutions! How to measure cultural change within a diverse institution?

We are delighted to present Salford’s ESS instructional case study which forms part of the Advance HE “inclusive institutions strand” of the Collaborative Development Fund 2021: New challenges, new solutions. The project enabled us to develop a deep understanding of the key cultural institutional enablers that drove the change required to facilitate ESS. Given the complexity associated with whole institutional change we gained a better understanding of the nuances in adopting and implementing a large institutional change project. The interactive case study developed for the Advance HE Project provides a narrative for how Salford culture articulated the enablers for transformational change to deliver the ESS framework. This presentation presents the array of technologies used in the data analysis and formulation of the interactive case study

Phase II pragmatic randomized controlled trial of patient-led therapies mMirror therapy and lower-limb exercises) during inpatient stroke rehabilitation

© American Society of Neurorehabilitation. Background and objective. Patient-led therapy has the potential to increase the amount of therapy patients undertake during stroke rehabilitation and to enhance recovery. Our objective was to assess the feasibility and acceptability of 2 patient-led therapies during the acute stages of stroke care: mirror therapy for the upper limb and lower-limb exercises for the lower limb. Methods. This was a blind assessed, multicenter, pragmatic randomized controlled trial of patient-led upper-limb mirror therapy and patient-led lower leg exercises. Stroke survivors with upper and lower limb limitations, undergoing inpatient rehabilitation and able to consent were recruited at least 1 week poststroke. Results. Both interventions proved feasible, with >90% retention. No serious adverse events were reported. Both groups did less therapy than recommended; typically 5 to 15 minutes for 7 days or less. Participants receiving mirror therapy (n = 63) tended to do less practice than those doing lower-limb exercises (n = 31). Those with neglect did 69% less mirror therapy than those without (P =.02), which was not observed in the exercise group. Observed between-group differences were modest but neglect, upper-limb strength, and dexterity showed some improvement in the mirror therapy group. No changes were seen in the lower-limb group. Conclusions. Both patient-led mirror therapy and lower-limb exercises during inpatient stroke care are safe, feasible, and acceptable and warrant further investigation. Practice for 5 to 15 minutes for 7 days is a realistic prescription unless strategies to enhance adherence are included