Mutations in ion channels involved in the generation and termination of action potentials constitute a family of molecular defects that underlie fatal cardiac arrhythmias in inherited long-QT syndrome. We report here that a loss-of-function (E1425G) mutation in ankyrin-B (also known as ankyrin 2), a member of a family of versatile membrane adapters, causes dominantly inherited type 4 long-QT cardiac arrhythmia in humans. Mice heterozygous for a null mutation in ankyrin-B are haploinsufficient and display arrhythmia similar to humans. Mutation of ankyrin-B results in disruption in the cellular organization of the sodium pump, the sodium/calcium exchanger, and inositol-1,4,5-trisphosphate receptors (all ankyrin-B-binding proteins), which reduces the targeting of these proteins to the transverse tubules as well as reducing overall protein level. Ankyrin-B mutation also leads to altered Ca2+ signalling in adult cardiomyocytes that results in extrasystoles, and provides a rationale for the arrhythmia. Thus, we identify a new mechanism for cardiac arrhythmia due to abnormal coordination of multiple functionally related ion channels and transporters.

Shoot branching is inhibited by auxin transported down the stem from the shoot apex. Auxin does not accumulate in inhibited buds and so must act indirectly. We show that mutations in the MAX4 gene of Arabidopsis result in increased and auxin-resistant bud growth. Increased branching in max4 shoots is restored to wild type by grafting to wild-type rootstocks, suggesting that MAX4 is required to produce a mobile branch-inhibiting signal, acting downstream of auxin. A similar role has been proposed for the pea gene, RMS1. Accordingly, MAX4 and RMS1 were found to encode orthologous, auxin-inducible members of the polyene dioxygenase family.Supplemental material is available at http://www.genesdev. org.Received December 6, 2002; revised version accepted March 20, 2003. Variation in shoot branching is an important cause of diversity in plant form. Individual species have a characteristic branching pattern, which can change through the life cycle in response to developmental cues and to environmental conditions (Cline 1991;Beveridge et al. 2003). Branching control therefore requires the integration of many signals, both known and unknown.Shoot branches arise from axillary meristems that form in the axils of leaves on the primary shoot axis. The axillary meristems themselves initiate leaves to form a bud. Bud growth can arrest but has the potential to reactivate to produce a shoot branch. Removal of the primary shoot apex results in activation of arrested axillary buds. The ability of the shoot apex to repress axillary bud growth is termed apical dominance. Thimann and Skoog (1933) reported that a compound, derived from the shoot apex, and later identified as auxin (indole-3-acetic acid), could inhibit the growth of lateral buds when applied to the stump of a decapitated plant. Subsequent work has provided multiple lines of evidence in support of auxinmediated bud inhibition in planta. However, a second messenger must relay the auxin signal into the bud because apically derived auxin is not transported into buds (Morris 1977) and exogenous auxin applied directly to buds does not inhibit their growth (Cline 1996).One model proposes that the effect of auxin on bud growth is mediated by cytokinin. Cytokinin can directly promote bud growth (Cline 1991); transgenic plants with increased auxin levels have reduced cytokinin levels (Eklö f et al. 2000), and cytokinin export from roots increases after decapitation, with this increase being abolished by application of auxin to the decapitated stump (Bangerth 1994). However, there is also good evidence for novel regulators of bud growth downstream of auxin. The ramosus mutants (rms1 to rms5) of pea (for reviews, see Beveridge 2000; Beveridge et al. 2003) have increased lateral branching, but this phenotype can be almost completely rescued by grafting a wild-type (WT) rootstock to an rms1, rms2, or rms5 mutant scion. Such grafting studies show that RMS1 and RMS5 are required for the production of a graft transmissible signal that moves from root to shoot and inhibits branching ...

Physiological and genetic studies with the ramosus (rms) mutants in garden pea (Pisum sativum) and more axillary shoots (max) mutants in Arabidopsis (Arabidopsis thaliana) have shown that shoot branching is regulated by a network of long-distance signals. Orthologous genes RMS1 and MAX4 control the synthesis of a novel graft-transmissible branching signal that may be a carotenoid derivative and acts as a branching inhibitor. In this study, we demonstrate further conservation of the branching control system by showing that MAX2 and MAX3 are orthologous to RMS4 and RMS5, respectively. This is consistent with the longstanding hypothesis that branching in pea is regulated by a novel long-distance signal produced by RMS1 and RMS5 and that RMS4 is implicated in the response to this signal. We examine RMS5 expression and show that it is more highly expressed relative to RMS1, but under similar transcriptional regulation as RMS1. Further expression studies support the hypothesis that RMS4 functions in shoot and rootstock and participates in the feedback regulation of RMS1 and RMS5 expression. This feedback involves a second novel long-distance signal that is lacking in rms2 mutants. RMS1 and RMS5 are also independently regulated by indole-3-acetic acid. RMS1, rather than RMS5, appears to be a key regulator of the branching inhibitor. This study presents new interactions between RMS genes and provides further evidence toward the ongoing elucidation of a model of axillary bud outgrowth in pea.

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Establishment and maintenance of transgenic mouse strains require being able to distinguish homozygous from heterozygous animals. To date, the developed real-time quantitative PCR techniques are often complicated, time-consuming and expensive. Here, we propose a very easy and rapid method with a simple data analysis to determine zygosity in transgenic mice. We show that the real-time quantitative PCR using SYBR Green fluorescent dye can be applied to discriminate two-fold differences in copy numbers of the transgene. Our procedure has to fit only three simple requirements: (1) to design primers capable of detecting one Ct difference for two-fold differences in DNA amounts (2) to measure genomic DNA concentrations accurately and (3) to have a reference animal of known zygosity in each run. Then, if the Ct values for the control gene are similar in all samples, we are able to compare directly the Ct values for the transgene in every sample, and so, to deduce the zygosity status of each mouse relative to the reference animal. This method is really simple and reliable, and it may be valuable as a rapid screening tool for zygosity status in transgenic animals.

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