The Leucine Zipper Domains of the Transcription Factors GCN4 and c-Jun Have Ribonuclease Activity

Basic-region leucine zipper (bZIP) proteins are one of the largest transcription factor families that regulate a wide range of cellular functions. Owing to the stability of their coiled coil structure leucine zipper (LZ) domains of bZIP factors are widely employed as dimerization motifs in protein engineering studies. In the course of one such study, the X-ray structure of the retro-version of the LZ moiety of yeast transcriptional activator GCN4 suggested that this retro-LZ may have ribonuclease activity. Here we show that not only the retro-LZ but also the authentic LZ of GCN4 has weak but distinct ribonuclease activity. The observed cleavage of RNA is unspecific, it is not suppressed by the ribonuclease A inhibitor RNasin and involves the breakage of 3′,5′-phosphodiester bonds with formation of 2′,3′-cyclic phosphates as the final products as demonstrated by HPLC/electrospray ionization mass spectrometry. Several mutants of the GCN4 leucine zipper are catalytically inactive, providing important negative controls and unequivocally associating the enzymatic activity with the peptide under study. The leucine zipper moiety of the human factor c-Jun as well as the entire c-Jun protein are also shown to catalyze degradation of RNA. The presented data, which was obtained in the test-tube experiments, adds GCN4 and c-Jun to the pool of proteins with multiple functions (also known as moonlighting proteins). If expressed in vivo, the endoribonuclease activity of these bZIP-containing factors may represent a direct coupling between transcription activation and controlled RNA turnover. As an additional result of this work, the retro-leucine zipper of GCN4 can be added to the list of functional retro-peptides

Divergent Cortical Generators of MEG and EEG during Human Sleep Spindles Suggested by Distributed Source Modeling

Background: Sleep spindles are,1-second bursts of 10–15 Hz activity, occurring during normal stage 2 sleep. In animals, sleep spindles can be synchronous across multiple cortical and thalamic locations, suggesting a distributed stable phaselocked generating system. The high synchrony of spindles across scalp EEG sites suggests that this may also be true in humans. However, prior MEG studies suggest multiple and varying generators. Methodology/Principal Findings: We recorded 306 channels of MEG simultaneously with 60 channels of EEG during naturally occurring spindles of stage 2 sleep in 7 healthy subjects. High-resolution structural MRI was obtained in each subject, to define the shells for a boundary element forward solution and to reconstruct the cortex providing the solution space for a noise-normalized minimum norm source estimation procedure. Integrated across the entire duration of all spindles, sources estimated from EEG and MEG are similar, diffuse and widespread, including all lobes from both hemispheres. However, the locations, phase and amplitude of sources simultaneously estimated from MEG versus EEG are highly distinct during the same spindles. Specifically, the sources estimated from EEG are highly synchronous across the cortex, whereas those from MEG rapidly shift in phase, hemisphere, and the location within the hemisphere. Conclusions/Significance: The heterogeneity of MEG sources implies that multiple generators are active during huma

Nouvas Rimas / da G. F. Caderas

NOUVAS RIMAS / DA G. F. CADERAS Nouvas Rimas / da G. F. Caderas (1) Einband (1) Titelseite (2) Engiadina (3) Uras greivas (19) Traducziuns (29) Föglias d'utuon (47) Algordaunzas (56) Register (68

Design, synthesis, and characterization of HIV-1 enhancer-binding polypeptides derived from bacteriophage 434 repressor

We have designed and synthesized HlV-1 enhancer-binding polypeptides that were derived from bacteriophage 434 repressor. These peptides were 39-54 residues long and contained either the recognition helix or the entire helix-turn-helix motif of the DNA-binding domain of 434 repressor. The dissociation constant of the complex formed between the standard peptide (R42) and a synthetic 70-bp HIV enhancer DNA was ca. 10-8 M. The specificity of the interaction of R42 with the two HIV enhancers was demonstrated by competitive band shift assays, stepwise displacement of the p50 subunit of transcription factor NF-κB from its two HIV enhancer binding sites, and DNase I footprinting; R42 seemed to protect best the two TTTCC sequences of the HIV enhancers against digestion by DNase I. R42 analogues with mutated recognition helix had lower DNA binding specificity. It remains to be investigated whether our artificial HIV enhancer-binding polypeptides are active in vivo