Studies of the response of peach and nectarine plants to gibberellin biosynthesis inhibitors in a hydroponic system

In small rooted peach and nectarine plants grown in a hydroponic system, low paclobutrazol (PBZ) levels applied to the roots suppressed shoot and root development, but to a different degree. A much stronger retarding effect was observed on the shoot, with a very limited effect on roots, resulting in a reduced shoot:root ratio. The effect of the inhibitor on the roots was a rapid increase in root diameter, with increased root branching upon recovery from the inhibiting effect. Root thickening, a typical response to PBZ, was detected also in nectarines when only the top was treated with the inhibitor, indicating a basal movement of PBZ, thus contradicting the accepted notion that its translocation is only acropetal. Root thickening was the earliest detected morphological response to PBZ, being observed already 3 days after exposure to the retardant. The return to a normal diameter was abrupt. Uniconazol (UNI) had a much stronger retarding effect on peach plants than did similar concentrations of PBZ. Reduced top growth resulted in a reduction in water consumption.Abbreviations PBZ paclobutrazol - UNI uniconazol - GA Gibberellin - GBI gibberellin biosynthesis inhibitor - ABA abscisic acid This research was supported by Grant no. 1-779-84 from the US-Israel Binational Agricultural Research and Development Fund (BARD).Contribution from the Agricultural research organization, The Volcani Center, Bet Dagan, Israel. No. 1506-E, 1994 series.     

Parental Somatic Mosaicism Is Underrecognized and Influences Recurrence Risk of Genomic Disorders

New human mutations are thought to originate in germ cells, thus making a recurrence of the same mutation in a sibling exceedingly rare. However, increasing sensitivity of genomic technologies has anecdotally revealed mosaicism for mutations in somatic tissues of apparently healthy parents. Such somatically mosaic parents might also have germline mosaicism that can potentially cause unexpected intergenerational recurrences. Here, we show that somatic mosaicism for transmitted mutations among parents of children with simplex genetic disease is more common than currently appreciated. Using the sensitivity of individual-specific breakpoint PCR, we prospectively screened 100 families with children affected by genomic disorders due to rare deletion copy-number variants (CNVs) determined to be de novo by clinical analysis of parental DNA. Surprisingly, we identified four cases of low-level somatic mosaicism for the transmitted CNV in DNA isolated from parental blood. Integrated probabilistic modeling of gametogenesis developed in response to our observations predicts that mutations in parental blood increase recurrence risk substantially more than parental mutations confined to the germline. Moreover, despite the fact that maternally transmitted mutations are the minority of alleles, our model suggests that sexual dimorphisms in gametogenesis result in a greater proportion of somatically mosaic transmitting mothers who are thus at increased risk of recurrence. Therefore, somatic mosaicism together with sexual differences in gametogenesis might explain a considerable fraction of unexpected recurrences of X-linked recessive disease. Overall, our results underscore an important role for somatic mosaicism and mitotic replicative mutational mechanisms in transmission genetics.     

The Butyrylcholinesterase K Variant Confers Structurally Derived Risks for Alzheimer Pathology?

The K variant of butyrylcholinesterase (BChE-K, 20% incidence) is a long debated risk factor for Alzheimer disease (AD). The A539T substitution in BChE-K is located at the C terminus, which is essential both for BChE tetramerization and for its capacity to attenuate β-amyloid (Aβ) fibril formation. Here, we report that BChE-K is inherently unstable as compared with the “usual” BChE (BChE-U), resulting in reduced hydrolytic activity and predicting prolonged acetylcholine maintenance and protection from AD. A synthetic peptide derived from the C terminus of BChE-K (BSP-K), which displayed impaired intermolecular interactions, was less potent in suppressing Aβ oligomerization than its BSP-U counterpart. Correspondingly, highly purified recombinant human rBChE-U monomers suppressed β-amyloid fibril formation less effectively than dimers, which also protected cultured neuroblastoma cells from Aβ neurotoxicity. Dual activity structurally derived changes due to the A539T substitution can thus account for both neuroprotective characteristics caused by sustained acetylcholine levels and elevated AD risk due to inefficient interference with amyloidogenic processes.Butyrylcholinesterase (BChE),³ the secondary acetylcholine (ACh)-hydrolyzing enzyme, is associated with the neurofibrillary tangles and amyloid plaques characteristic of Alzheimer disease (AD) (1), which suggests that it functions as a potential AD modulator. BChE activity increases in the AD brain (2–4), where it co-localizes with β-amyloid (Aβ) fibrils (5, 6). Aβ is a 39–42-amino-acid amphiphilic peptide, derived from the transmembrane domain and extracellular region of the Aβ precursor protein (7). At high concentrations, Aβ acquires a β-sheet structure, becomes insoluble, and accumulates in neurotoxic oligomers and fibrils (8) to become the main constituent of plaques in the brain of AD patients. Recent hypotheses attribute causal roles in AD to presenilin (9), oxidative stress (10), metals (11), double hit origin (12), or mitochondrial damage (13). The alternative theories state that Aβ represents a bystander or even a protector rather than the causative factor of disease and that Aβ amyloidogenesis is secondary to other pathogenic events (14). Nevertheless, a wealth of evidence demonstrates a pivotal role for Aβ in the pathogenesis of AD, yielding the amyloid cascade hypothesis (15). According to this hypothesis, the pathological accumulation of Aβ in the brain leads to oxidative stress, neuronal destruction, and finally, the clinical syndrome of AD. It is within this context that we have studied the interactions of the Kalow variant (BChE-K) with Aβ.The C terminus of BChE functions as a tetramerization domain (16, 17) and is responsible for its quaternary organization. Four BChE monomers are held together by the aromatic interactions of seven highly conserved aromatic residues, termed the tryptophan amphiphilic tetramerization domain (WAT) (16, 17). The WAT domain interacts with proline-rich attachment domains, either via proline-rich membrane anchor in brain neurons (18) or, in neuromuscular junctions, with cholinesterase-associated collagen Q (19). In the serum, BChE tetramerization is supported by an analogous 17-mer proline-rich peptide derived from lamellipodin (20).Analyzing the quaternary organization of cholinesterases is a complicated task. To date, all biologically relevant crystal structures of cholinesterases have been truncated forms that lack the C terminus of the protein (21), apart from a more recent study of full-length BChE that yielded crystal packing, which did not allow C-terminal interactions among subunits and lacked electron densities in the C terminus region, indicating structural disorder Protein Data Bank (PDB) code 1VZJ (22). Of note, the crystal structure of the homologous C terminus of tetrameric synaptic acetylcholinesterase (AChE-S) could only be determined based on synthetic peptides derived from the sequence of the AChE-S tail and stabilized with a proline-rich attachment domain (23).In addition to the “usual” (BChE-U) form, BChE has nearly 40 genomic variants. The most common is BChE-K, with allelic frequencies of 0.13–0.21. BChE-K includes a single nucleotide polymorphism at position 1699 (single nucleotide polymorphism data base (dbSNP) ID: rs1803274; alleles, A/G). This leads to an alanine-to-threonine substitution at position 539, 36 residues upstream to the C terminus of BChE (24), within the tetramerization domain that we previously found to attenuate amyloid fibril formation (25).Ample evidence supports the importance of alanine-to-threonine substitutions and their relevance to amyloidogenic processes, protein stability, and quaternary organization (supplemental Table ST1). Point mutations at the dimer interface of light chain immunoglobulins decrease their stability so that the A34T polymorphism in this protein leads to systemic amyloidosis (26). An A25T mutant of the tetrameric human protein Transthyretin (TTR), associated with central nervous system amyloidosis, is prone to aggregation and exhibits drastically reduced tertiary and quaternary structural stabilities (27). The thermodynamic stability profile of the A25T TTR mutant shows that both monomers and tetramers of this variant are highly destabilized. In addition, A25T TTR tetramers dissociate very rapidly (about 1200-fold faster than the dissociation of wild-type TTR), reflecting a high degree of kinetic destabilization of their quaternary structure. These factors together probably contribute to the high propensity of A25T TTR to aggregate in vitro.The capacity of serum BChE-K to hydrolyze butyrylthiocholine was reported to be reduced by 30% relative to BChE-U, for yet unclear reasons (24). The reduced hydrolytic activity of BChE-K predicts that BChE-K carriers would potentially sustain improved cholinergic transmission as compared with BChE-U carriers and has been shown to correlate with preserved performance of attention and reduced rates of cognitive decline (28). However, BChE-K carriers are refractory to cholinesterase inhibitor therapy, the current leading treatment of AD (29). This raised the question whether BChE-K functions as an AD risk or protection factor. Genotype studies are controversial, with some showing increased risk of AD for homozygote BChE-K carriers (e.g. Ref. 30), whereas others suggest a protective effect (e.g. Ref. 31). A recent meta-analysis concluded that on average, BChE-K is neither a risk factor nor a protection factor for AD (32). Based on our previous findings of the arrest of Aβ fibril formation by BChE and considering the accumulation of monomeric BChE in the most severe AD cases (33), we used a variety of chemical techniques to study the effect of the A539T substitution on BChE stability and tetramerization on the one hand and on its potency in attenuating Aβ oligomerization and fibril formation on the other.     

CXCR7 Functions as a Scavenger for CXCL12 and CXCL11

Background

CXCR7 (RDC1), the recently discovered second receptor for CXCL12, is phylogenetically closely related to chemokine receptors, but fails to couple to G-proteins and to induce typical chemokine receptor mediated cellular responses. The function of CXCR7 is controversial. Some studies suggest a signaling activity in mammalian cells and zebrafish embryos, while others indicate a decoy activity in fish. Here we investigated the two propositions in human tissues.

Methodology/Principal Findings

We provide evidence and mechanistic insight that CXCR7 acts as specific scavenger for CXCL12 and CXCL11 mediating effective ligand internalization and targeting of the chemokine cargo for degradation. Consistently, CXCR7 continuously cycles between the plasma membrane and intracellular compartments in the absence and presence of ligand, both in mammalian cells and in zebrafish. In accordance with the proposed activity as a scavenger receptor CXCR7-dependent chemokine degradation does not become saturated with increasing ligand concentrations. Active CXCL12 sequestration by CXCR7 is demonstrated in adult mouse heart valves and human umbilical vein endothelium.

Conclusions/Significance

The finding that CXCR7 specifically scavenges CXCL12 suggests a critical function of the receptor in modulating the activity of the ubiquitously expressed CXCR4 in development and tumor formation. Scavenger activity of CXCR7 might also be important for the fine tuning of the mobility of hematopoietic cells in the bone marrow and lymphoid organs.     

Dynamics of protein distributions in cell populations

A population of cells exhibits wide phenotypic variation even if it is genetically homogeneous. In particular, individual cells differ from one another in the amount of protein they express under a given regulatory system under fixed conditions. Here we study how protein distributions in a population of the yeast S. cerevisiae are shaped by a balance of processes: protein production--an intracellular process--and protein dilution due to cell division--a population process. We measure protein distributions by employing reporter green fluorescence protein (gfp) under the regulation of the yeast GAL system under conditions where it is metabolically essential. Cell populations are grown in chemostats, thus allowing control of the environment and stable measurements of distribution dynamics over many generations. Despite the essential functional role of the GAL system in a pure galactose medium, steady-state distributions are found to be universally broad, with exponential tails and a large standard-deviation-to-mean ratio. Under several different perturbations the dynamics of the distribution is observed to be asymmetric, with a much longer time to build a wide expression distribution from below compared with a fast relaxation of the distribution toward steady state from above. These results show that the main features of the protein distributions are largely determined by population effects and are less sensitive to the intracellular biochemical noise.     

Found in translation: A new player in EMT

Epithelial mesenchymal transition (EMT) is a complex process that involves changes in gene expression, cytoskeleton organization, cell adhesion, and extracellular matrix composition. Screening for genes mediating EMT and cancer metastasis, Waerner, Alacakaptan, and colleagues identified ILEI, a cytokine-like protein that plays an essential role in EMT, tumor growth, and late steps of metastasis.     

Prenylation-deficient G protein gamma subunits disrupt GPCR signaling in the zebrafish

Prenylation of G protein gamma (γ) subunits is necessary for the membrane localization of heterotrimeric G proteins and for functional heterotrimeric G protein coupled receptor (GPCR) signaling. To evaluate GPCR signaling pathways during development, we injected zebrafish embryos with mRNAs encoding Gγ subunits mutated so that they can no longer be prenylated. Low-level expression of these prenylation-deficient Gγ subunits driven either ubiquitously or specifically in the primordial germ cells (PGCs) disrupts GPCR signaling and manifests as a PGC migration defect. This disruption results in a reduction of calcium accumulation in the protrusions of migrating PGCs and a failure of PGCs to directionally migrate. When co-expressed with a prenylation-deficient Gγ, 8 of the 17 wildtype Gγ isoforms individually confer the ability to restore calcium accumulation and directional migration. These results suggest that while the Gγ subunits possess the ability to interact with G Beta (β) proteins, only a subset of wildtype Gγ proteins are stable within PGCs and can interact with key signaling components necessary for PGC migration. This in vivo study highlights the functional redundancy of these signaling components and demonstrates that prenylation-deficient Gγ subunits are an effective tool to investigate the roles of GPCR signaling events during vertebrate development.