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Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) that compromise its chloride channel activity. The most common mutation, p.Phe508del, results in the production of a misfolded CFTR protein, which has residual channel activity but is prematurely degraded. Because of the inherent complexity of the pathogenetic mechanisms involved in CF, which include impaired chloride permeability and persistent lung inflammation, a multidrug approach is required for efficacious CF therapy. To date, no individual drug with pleiotropic beneficial effects is available for CF. Here we report on the ability of thymosin alpha 1 (Tα1)—a naturally occurring polypeptide with an excellent safety profile in the clinic when used as an adjuvant or an immunotherapeutic agent—to rectify the multiple tissue defects in mice with CF as well as in cells from subjects with the p.Phe508del mutation. Tα1 displayed two combined properties that favorably opposed CF symptomatology: it reduced inflammation and increased CFTR maturation, stability and activity. By virtue of this two-pronged action, Tα1 has strong potential to be an efficacious single-molecule-based therapeutic agent for CF. more...

1The inwardly rectifying potassium channel Kir5.1 appears to form functional channels only by coexpression with either Kir4.1 or Kir4.2. Kir4.1‐Kir5.1 heteromeric channels have been shown to exist in vivoin renal tubular epithelia. However, Kir5.1 is expressed in many other tissues where Kir4.1 is not found. Using Kir5.1‐specific antibodies we have localised Kir5.1 expression in the pancreas, a tissue where Kir4.2 is also highly expressed.2Heteromeric Kir5.1‐Kir4.1 channels are significantly more sensitive to intracellular acidification than Kir4.1 currents. We demonstrate that this increased sensitivity is primarily due to modulation of the intrinsic Kir4.1 pH sensitivity by Kir5.1.3Kir4.2 was found to be significantly more pH sensitive (pKa= 7.1) than Kir4.1 (pKa= 5.99) due to an additional pH‐sensing mechanism involving the C‐terminus. As a result, coexpression with Kir5.1 does not cause a major shift in the pH sensitivity of the heteromeric Kir4.2‐Kir5.1 channel.4Cell‐attached single channel analysis of Kir4.2 revealed a channel with a high open probability (Po> 0.9) and single channel conductance of ˜25 pS, whilst coexpression with Kir5.1 produced novel bursting channels (Po< 0.3) and a principal conductance of ˜54 pS with several subconductance states.5These results indicate that Kir5.1 may form heteromeric channels with Kir4.2 in tissues where Kir4.1 is not expressed (e.g. pancreas) and that these novel channels are likely to be regulated by changes in intracellular pH. In addition, the extreme pH sensitivity of Kir4.2 has implications for the role of this subunit as a homotetrameric channel. more...

The physiological role of the inwardly rectifying potassium channel, Kir5.1, is poorly understood, as is the molecular identity of many renal potassium channels. In this study we have used Kir5.1-specific antibodies to reveal abundant expression of Kir5.1 in renal tubular epithelial cells, where Kir4.1 is also expressed. Moreover, we also show that Kir5.1/Kir4.1 heteromeric channel activity is extremely sensitive to inhibition by intracellular acidification and that this novel property is conferred predominantly by the Kir5.1 subunit. These findings suggest that Kir5.1/Kir4.1 heteromeric channels are likely to exist in vivoand implicate an important and novel functional role for the Kir5.1 subunit. more...

Episodic ataxia type‐1 syndrome (EA‐1) is an autosomal dominant neurological disorder that manifests itself during infancy and results from point mutations in the voltage‐gated potassium channel gene hKv1.1. The hallmark of the disease is continuous myokymia and episodic attacks of spastic contractions of the skeletal muscles, which cause permanent disability. Coexpression of hKv1.1 and hKv1.2 subunits produces heteromeric potassium channels with biophysical and pharmacological properties intermediate between the respective homomers. By using tandemly linked subunits, we demonstrate that hKv1.1 subunits bearing the EA‐1 mutations V408A and E325D combine with hKv1.2 to produce channels with altered kinetics of activation, deactivation, C‐type inactivation, and voltage dependence. Moreover, hKv1.1V408A single‐channel analysis reveals a ∼threefold reduction of the mean open duration of the channel compared with the wild‐type, and this mutation alters the open‐state stability of both homomeric and heteromeric channels. The results demonstrate that human Kv1.2 and Kv1.1 subunits coassemble to form a novel channel with distinct gating properties that are altered profoundly by EA‐1 mutations, thus uncovering novel physiopathogenetic mechanisms of episodic ataxia type‐1 myokymia syndrome.—D'Adamo, M. C., Imbrici, P., Sponcichetti, F., Pessia, M. Mutations in the KCNA1gene associated with episodic ataxia type‐1 syndrome impair heteromeric voltage‐gated K+channel function. FASEB J.13, 1335–1345 (1999) more...

Kir 5.1 is a member of the inward rectifier potassium channel superfamily which does not form functional channels when expressed by itself in Xenopus laevisoocytes. rt‐PCR reveals high levels of Kir 5.1 mRNA expression in testis but the function of this channel remains unknown. To determine the cell‐specific expression of this channel in the testis we raised a polyclonal antibody against an external epitope of Kir 5.1 and tested its specificity in Xenopusoocytes expressing several cloned Kir subunits. Strong immunoreactivity for Kir 5.1 was found in seminiferous tubules of rat testis and, particularly, in spermatogonia, primary and secondary spermatocytes, spermatids and in the head and body of spermatozoa. The intensity of Kir 5.1 immunofluorescence, quantified using laser scanning microscopy, increased with age at every stage in the development of sperm from spermatogonia and reached a peak in 60‐day‐old rats. In contrast, the immunofluorescence decreased in 90‐day‐old animals and was detected mostly in spermatozoa. The results demonstrate that Kir 5.1 expression in the testis is localised to cells involved in spermatogenesis, showing a temporal pattern of expression during sexual maturity. more...

In the version of this article originally published, the amino acid sequence for Tα1 described in the Online Methods is incorrect. The sequence is described as “Ac-SDAAVDTSSEITTJDLKEKKEVVEEAEN-OH”. It should be “Ac-SDAAVDTSSEITTKDLKEKKEVVEEAEN-OH”. The error has been corrected in the HTML and PDF versions of this article. more...

In the version of this article originally published, some labels in Fig. 1f are incorrect. The “β-actin” labels on the second and fourth rows of blots should instead be “β-tubulin”. The error has been corrected in the HTML and PDF versions of this article. more...