Steven E. Mansoor

Consensus and Variant cAMP-regulated Enhancers Have Distinct CREB-binding Properties

Recent determination of the cAMP response element-binding protein (CREB) basic leucine zipper (bZIP) consensus CRE crystal structure revealed key dimerization and DNA binding features that are conserved among members of the CREB/CREM/ATF-1 family of transcription factors. Dimerization appeared to be mediated by a Tyr307–Glu312interhelical hydrogen bond and a Glu319–Arg314electrostatic interaction. An unexpected hexahydrated Mg2+ion was centered above the CRE in the dimer cavity. In the present study, we related these features to CREB dimerization and DNA binding. A Y307F substitution reduced dimer stability and DNA binding affinity, whereas a Y307R mutation produced a stabilizing effect. Mutation of Glu319 to Ala or Lys attenuated dimerization and DNA binding. Mg2+ ions enhanced the binding affinity of wild-type CREB to the palindromic CRE by ∼20-fold but did not do so for divergent CREs. Similarly, mutation of Lys304, which mediates the CREB interaction with the hydrated Mg2+, blocked CREB binding to the palindromic but not the variant CRE sequences. The distinct binding characteristics of the K304A mutants to the consensus and variant CRE sequences indicate that CREB binding to these elements is differentially regulated by Mg2+ ions. We suggest that CREB binds the consensus and variant CRE sequences through fundamentally distinct mechanisms.

X-ray structures define human P2X3 receptor gating cycle and antagonist action.

P2X receptors are trimeric, non-selective cation channels activated by ATP that have important roles in the cardiovascular, neuronal and immune systems. Despite their central function in human physiology and although they are potential targets of therapeutic agents, there are no structures of human P2X receptors. The mechanisms of receptor desensitization and ion permeation, principles of antagonism, and complete structures of the pore-forming transmembrane domains of these receptors remain unclear. Here we report X-ray crystal structures of the human P2X3 receptor in apo/resting, agonist-bound/open-pore, agonist-bound/closed-pore/desensitized and antagonist-bound/closed states. The open state structure harbours an intracellular motif we term the ‘cytoplasmic cap’, which stabilizes the open state of the ion channel pore and creates lateral, phospholipid-lined cytoplasmic fenestrations for water and ion egress. The competitive antagonists TNP-ATP and A-317491 stabilize the apo/resting state and reveal the interactions responsible for competitive inhibition. These structures illuminate the conformational rearrangements that underlie P2X receptor gating and provide a foundation for the development of new pharmacological agents.

IMAGES IN CLINICAL MEDICINE. Lancisi's Sign

A 60-year-old man presented with progressive dyspnea and weight gain. A holosystolic murmur that augmented with inspiration was noted at the left lower sternal border. Examination of the neck revealed a palpable, monomorphic venous pulsation, known as Lancisis sign, shown in a video.

Quincke's pulse

A 34-year-old man presented with a 6-to-8-week history of fatigue, malaise, fevers, and night sweats. He had a history of a congenital quadricuspid aortic valve and 16 years earlier had undergone a Ross procedure.