Endogenous N-terminal Domain Cleavage Modulates α1D-Adrenergic Receptor Pharmacodynamics.

The α1D-adrenergic receptor (ADRA1D) is a key regulator of cardiovascular, prostate, and central nervous system functions. This clinically relevant G protein-coupled receptor has proven difficult to study, as it must form an obligate modular homodimer containing the PDZ proteins scribble and syntrophin or become retained in the endoplasmic reticulum as non-functional protein. We previously determined that targeted removal of the N-terminal (NT) 79 amino acids facilitates ADRA1D plasma membrane expression and agonist-stimulated functional responses. However, whether such an event occurs in physiological contexts was unknown. Herein, we report the ADRA1D is subjected to innate NT processing in cultured human cells. SNAP near-infrared imaging and tandem-affinity purification revealed the ADRA1D is expressed as both full-length and NT truncated forms in multiple human cell lines. Serial truncation mapping identified the cleavage site as Leu(90)/Val(91) in the 95-amino acid ADRA1D NT domain, suggesting human cells express a Δ1-91 ADRA1D species. Tandem-affinity purification MS/MS and co-immunoprecipitation analysis indicate NT processing of ADRA1D is not required to form scribble-syntrophin macromolecular complexes. Yet, label-free dynamic mass redistribution signaling assays demonstrate that Δ1-91 ADRA1D agonist responses were greater than WT ADRA1D. Mutagenesis of the cleavage site nullified the processing event, resulting in ADRA1D agonist responses less than the WT receptor. Thus, we propose that processing of the ADRA1D NT domain is a physiological mechanism employed by cells to generate a functional ADRA1D isoform with optimal pharmacodynamic properties.

Health care resource utilization associated with treatment of penicillin-susceptible and -nonsusceptible isolates of Streptococcus pneumoniae.

Despite failure to correlate in vitro susceptibility with clinical outcomes for respiratory tract infections and bacteremia, resistance affects management of patients with pneumococcal infections. The economic impact of resistance among pneumococci has not been evaluated. We conducted a single-center, retrospective, observational, cohort study of hospitalized patients infected with Streptococcus pneumoniae isolated from blood or a respiratory source between January 1, 1995, and December 31, 1998. Data were collected for 36 days surrounding the day that the first positive culture was collected. Patients were grouped according to isolate penicillin-susceptibility profile [susceptible minimum inhibitory concentration (MIC) < or = 0.06 microg/ml, nonsusceptible MIC > or = 0.125 microg/ml), and data were analyzed with respect to health care resource utilization patterns. Of 231 patients identified, 142 and 89 had susceptible and nonsusceptible isolates, respectively. Groups were similar with respect to demographics and comorbidities, except that patients infected with a nonsusceptible isolate were more likely to have the isolate obtained from a respiratory source and to have a history of recent antibiotic therapy. No difference was noted with respect to clinical outcome; however, patients infected with a nonsusceptible isolate had a longer median stay (14 vs 10 days, p<0.05). They also had significantly higher total median costs (1600 dollars, 95% confidence interval 257-2943 dollars) due to room and nursing services. Infections caused by penicillin-nonsusceptible pneumococci were not associated with a worse outcome in hospitalized patients but were associated with increased cost of care.