A Highlights from MBoC Selection: Forty-five years of cGMP research in Dictyostelium: understanding the regulation and function of the cGMP pathway for cell movement and chemotaxis

In Dictyostelium, chemoattractants induce a fast cGMP response that mediates myosin filament formation in the rear of the cell. The major cGMP signaling pathway consists of a soluble guanylyl cyclase sGC, a cGMP-stimulated cGMP-specific phosphodiesterase, and the cGMP-target protein GbpC. Here we combine published experiments with many unpublished experiments performed in the past 45 years on the regulation and function of the cGMP signaling pathway. The chemoattractants stimulate heterotrimeric Gαβγ and monomeric Ras proteins. A fraction of the soluble guanylyl cyclase sGC binds with high affinity to a limited number of membrane binding sites, which is essential for sGC to become activated by Ras and Gα proteins. sGC can also bind to F-actin; binding to branched F-actin in pseudopods enhances basal sGC activity, whereas binding to parallel F-actin in the cortex reduces sGC activity. The cGMP pathway mediates cell polarity by inhibiting the rear: in unstimulated cells by sGC activity in the branched F-actin of pseudopods, in a shallow gradient by stimulated cGMP formation in pseudopods at the leading edge, and during cAMP oscillation to erase the previous polarity and establish a new polarity axis that aligns with the direction of the passing cAMP wave.     

The Emerging Role of Copeptin

Direct measurement of the nonapeptide vasopressin has been limited by analyte instability ex vivo and in vivo rapid degradation, low serum concentrations requiring a sensitive assay and inherent secretory pulsatility. Copeptin is a 39 amino acid glycopeptide cleavage product of vasopressin synthesis with high stability, providing a marker of vasopressin secretion. Copeptin measurement has applications in diagnosis of diabetes insipidus and other diseases with altered vasopressin secretion. This review summarises our current understanding of serum copeptin measurement in diabetes insipidus and possible future applications of copeptin assays. As vasopressin is a stress hormone, there is emerging evidence on the use of copeptin for diagnosis and prognostication of disorders such as syndrome of inappropriate anti-diuretic hormone secretion, diabetes mellitus, critical illness, stroke, cardiovascular disease, respiratory disease, renal disease and thermal stress. Copeptin concentration measurement is likely to improve the diagnostic reliability of diabetes insipidus and, as a marker of stress, may have diagnostic or prognostic utility in specific clinical circumstances. Further studies are needed to determine if goal-directed therapy using plasma copeptin concentrations may improve patient outcomes.     

Activation of soluble guanylyl cyclase at the leading edge during Dictyostelium chemotaxis

Dictyostelium contains two guanylyl cyclases, GCA, a 12-transmembrane enzyme, and sGC, a homologue of mammalian soluble adenylyl cyclase. sGC provides nearly all chemoattractant-stimulated cGMP formation and is essential for efficient chemotaxis toward cAMP. We show that in resting cells the major fraction of the sGC-GFP fusion protein localizes to the cytosol, and a small fraction is associated to the cell cortex. With the artificial substrate Mn2+/GTP, sGC activity and protein exhibit a similar distribution between soluble and particulate fraction of cell lysates. However, with the physiological substrate Mg2+/GTP, sGC in the cytosol is nearly inactive, whereas the particulate enzyme shows high enzyme activity. Reconstitution experiments reveal that inactive cytosolic sGC acquires catalytic activity with Mg2+/GTP upon association to the membrane. Stimulation of cells with cAMP results in a twofold increase of membrane-localized sGC-GFP, which is accompanied by an increase of the membrane-associated guanylyl cyclase activity. In a cAMP gradient, sGC-GFP localizes to the anterior cell cortex, suggesting that in chemotacting cells, sGC is activated at the leading edge of the cell.     

Renal glutathione homeostasis in compensatory renal growth

Glutathione homeostasis was investigated in unilaterally nephrectomized and sham-operated rats. Following twelve days of compensatory renal growth, it was found that the concentrations of glutathione and glutathione disulfide in representative samples of the entire remnant right kidney from the nephrectomized rats were similar to those found in corresponding samples of the right kidneys from the sham-operated rats. However, since the mass of the remnant right kidneys in the nephrectomized rats was greater than that of the right kidneys from the sham-operated rats, the absolute content of glutathione and glutathione disulfide was greater in the remnant right kidneys of the nephrectomized rats than in the right kidneys of the sham-operated rats. In general, the findings from the present study indicate that the absolute content of glutathione and glutathione disulfide in renal epithelial cells increases in proportion to the increase in mass that results from compensatory renal cellular hypertrophy.     

Cyclical Action of the WASH Complex: FAM21 and Capping Protein Drive WASH Recycling,Not Initial Recruitment

1. Download : Download high-res image (181KB)
2. Download : Download full-size image

Highlights? Dictyostelium WASH complex neutralizes lysosomes, then is recycled in small vesicles ? FAM21 is required at a different step than the other four WASH complex subunits ? Mutants in FAM21 can make actin on vesicles but cannot recycle WASH complex after use ? WASH complex is recycled coupling between actin, capping protein, and FAM21     

Extraversion is linked to volume of the orbitofrontal cortex and amygdala

Neuroticism and extraversion are personality factors associated with the vulnerability for developing depression and anxiety disorders, and are possibly differentially related to brain structures implicated in the processing of emotional information and the generation of mood states. To date, studies on brain morphology mainly focused on neuroticism, a dimension primarily related to negative affect, yielding conflicting findings concerning the association with personality, partially due to methodological issues and variable population samples under study. Recently, extraversion, a dimension primarily related to positive affect, has been repeatedly inversely related to with symptoms of depression and anxiety disorders. In the present study, high resolution structural T1-weighted MR images of 65 healthy adults were processed using an optimized Voxel Based Morphometry (VBM) approach. Multiple regression analyses were performed to test for associations of neuroticism and extraversion with prefrontal and subcortical volumes. Orbitofrontal and right amygdala volume were both positively related to extraversion. Extraversion was differentially related to volume of the anterior cingulate cortex in males (positive) and females (negative). Neuroticism scores did not significantly correlate with these brain regions. As extraversion is regarded a protective factor for developing anxiety disorders and depression and has been related to the generation of positive affect, the present results indicate that the reduced likelihood of developing affective disorders in individuals high on extraversion is related to modulation of emotion processing through the orbitofrontal cortex and the amygdala.     

Exome sequencing identifies truncating mutations in human SERPINF1 in autosomal-recessive osteogenesis imperfecta

Osteogenesis imperfecta (OI) is a heterogeneous genetic disorder characterized by bone fragility and susceptibility to fractures after minimal trauma. After mutations in all known OI genes had been excluded by Sanger sequencing, we applied next-generation sequencing to analyze the exome of a single individual who has a severe form of the disease and whose parents are second cousins. A total of 26,922 variations from the human reference genome sequence were subjected to several filtering steps. In addition, we extracted the genotypes of all dbSNP130-annotated SNPs from the exome sequencing data and used these 299,494 genotypes as markers for the genome-wide identification of homozygous regions. A single homozygous truncating mutation, affecting SERPINF1 on chromosome 17p13.3, that was embedded into a homozygous stretch of 2.99 Mb remained. The mutation was also homozygous in the affected brother of the index patient. Subsequently, we identified homozygosity for two different truncating SERPINF1 mutations in two unrelated patients with OI and parental consanguinity. All four individuals with SERPINF1 mutations have severe OI. Fractures of long bones and severe vertebral compression fractures with resulting deformities were observed as early as the first year of life in these individuals. Collagen analyses with cultured dermal fibroblasts displayed no evidence for impaired collagen folding, posttranslational modification, or secretion. SERPINF1 encodes pigment epithelium-derived factor (PEDF), a secreted glycoprotein of the serpin superfamily. PEDF is a multifunctional protein and one of the strongest inhibitors of angiogenesis currently known in humans. Our data provide genetic evidence for PEDF involvement in human bone homeostasis.     

ANTHROPOMETRIC DETERMINANTS OF ROWING ERGOMETER PERFORMANCE IN PHYSICALLY INACTIVE COLLEGIATE FEMALES

The aim of the study was to evaluate anthropometric characteristics as determinants of 500 m rowing ergometer performance in physically inactive collegiate females. In this cross-sectional study, which included 196 collegiate females aged 19-23 years not participating in regular physical activities, body mass (BM), body height (BH), length of upper limbs (LA), length of lower limbs (LL), body mass index (BMI), slenderness index (SI), and the Choszcz-Podstawski index (CPI) were measured and a stepwise multiple regression analysis was performed. Participants performed 500 m maximal effort on a Concept II rowing ergometer. BM, BH, LA, LL, and the BMI, SI and CPI indices were found to be statistically significant determinants of 500 m performance. The best results (T) were achieved by females whose BH ranged from 170 to 180 cm, with LA and LL ranging from 75 to 80 cm and 85 to 90 cm, respectively. The best fitting statistical model was identified as: T = 11.6793 L_R – 0.1130 L_R² – 0.0589 L_N² + 29.2157 CPI² + 0.1370 LR·LN - 2.6926 LR·CPI – 211.7796. This study supports a need for additional studies focusing on understanding the importance of anthropometric differences in rowing ergometer performance, which could lead to establishing a better quality reference for evaluation of cardiorespiratory fitness tested using a rowing ergometer in collegiate females.