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Methods for early treatment response evaluation to systemic therapy of liver metastases are lacking. Tumor tissue often exhibits an increased ratio of phosphomonoesters to phosphodiesters (PME/PDE), which can be noninvasively measured by phosphorus magnetic resonance spectroscopy (31P MRS), and may be a marker for early therapy response assessment in liver metastases. However, with commonly used 31P surface coils for liver 31P MRS, the liver is not fully covered, and metastases may be missed. The objective of this study was to demonstrate the feasibility of 31P MRS imaging (31P MRSI) with full liver coverage to assess 31P metabolite levels and chemotherapy‐induced changes in liver metastases of gastro‐esophageal cancer, using a 31P whole‐body birdcage transmit coil in combination with a 31P body receive array at 7 T. 3D 31P MRSI data were acquired in two patients with hepatic metastases of esophageal cancer, before the start of chemotherapy and after 2 (and 9 in patient 2) weeks of chemotherapy. 3D 31P MRSI acquisitions were performed using an integrated 31P whole‐body transmit coil in combination with a 16‐channel body receive array at 7 T, with a field of view covering the full abdomen and a nominal voxel size of 20‐mm isotropic. From the 31P MRSI data, 12 31P metabolite signals were quantified. Prior to chemotherapy initiation, both PMEs, that is, phosphocholine (PC) and phosphoethanolamine (PE), were significantly higher in all metastases compared with the levels previously determined in the liver of healthy volunteers. After 2 weeks of chemotherapy, PC and PE levels remained high or even increased further, resulting in increased PME/PDE ratios compared with healthy liver tissue, in correspondence with the clinical assessment of progressive disease after 2 months of chemotherapy. The suggested approach may present a viable tool for early therapy (non)response assessment of tumor metabolism in patients with liver metastases. [ABSTRACT FROM AUTHOR]

Quantitative three‐dimensional (3D) imaging of phosphorus (31P) metabolites is potentially a promising technique with which to assess the progression of liver disease and monitor therapy response. However, 31P magnetic resonance spectroscopy has a low sensitivity and commonly used 31P surface coils do not provide full coverage of the liver. This study aimed to overcome these limitations by using a 31P whole‐body transmit coil in combination with a 16‐channel 31P receive array at 7 T. Using this setup, we determined the repeatability of whole‐liver 31P magnetic resonance spectroscopic imaging (31P MRSI) in healthy subjects and assessed the effects of principal component analysis (PCA)‐based denoising on the repeatability parameters. In addition, spatial variations of 31P metabolites within the liver were analyzed. 3D 31P MRSI data of the liver were acquired with a nominal voxel size of 20 mm isotropic in 10 healthy volunteers twice on the same day. Data were reconstructed without denoising, and with PCA‐based denoising before or after channel combination. From the test–retest data, repeatability parameters for metabolite level quantification were determined for 12 31P metabolite signals. On average, 31P MR spectra from 100 ± 25 voxels in the liver were analyzed. Only voxels with contamination from skeletal muscle or the gall bladder were excluded and no voxels were discarded based on (low) signal‐to‐noise ratio (SNR). Repeatability for most quantified 31P metabolite levels in the liver was good to excellent, with an intrasubject variability below 10%. PCA‐based denoising increased the SNR ~ 3‐fold, but did not improve the repeatability for mean liver 31P metabolite quantification with the fitting constraints used. Significant spatial heterogeneity of various 31P metabolite levels within the liver was observed, with marked differences for the phosphomonoester and phosphodiester metabolites between the left and right lobe. In conclusion, using a 31P whole‐body transmit coil in combination with a 16‐channel 31P receive array at 7 T allowed 31P MRSI acquisitions with full liver coverage and good to excellent repeatability. [ABSTRACT FROM AUTHOR]

The main inhibitory neurotransmitter receptors in the adult central nervous system (CNS) are type A γ-aminobutyric acid receptors (GABAARs) and glycine receptors (GlyRs). Synaptic responses mediated by GlyR and GABAAR display a hyperpolarizing shift during development. This shift relies mainly on the developmental up-regulation of the K+-Cl− co-transporter KCC2 responsible for the extrusion of Cl−. In mature neurons, altered KCC2 function—mainly through increased endocytosis—leads to the re-emergence of depolarizing GABAergic and glycinergic signaling, which promotes hyperexcitability and pathological activities. Identifying signaling pathways and molecular partners that control KCC2 surface stability thus represents a key step in the development of novel therapeutic strategies. Here, we present our current knowledge on the cellular and molecular mechanisms governing the plasma membrane turnover rate of the transporter under resting conditions and in response to synaptic activity. We also discuss the notion that KCC2 lateral diffusion is one of the first parameters modulating the transporter membrane stability, allowing for rapid adaptation of Cl− transport to changes in neuronal activity.

The main inhibitory neurotransmitter receptors in the adult central nervous system (CNS) are type A γ-aminobutyric acid receptors (GABAARs) and glycine receptors (GlyRs). Synaptic responses mediated by GlyR and GABAAR display a hyperpolarizing shift during development. This shift relies mainly on the developmental up-regulation of the K+-Cl− co-transporter KCC2 responsible for the extrusion of Cl−. In mature neurons, altered KCC2 function—mainly through increased endocytosis—leads to the re-emergence of depolarizing GABAergic and glycinergic signaling, which promotes hyperexcitability and pathological activities. Identifying signaling pathways and molecular partners that control KCC2 surface stability thus represents a key step in the development of novel therapeutic strategies. Here, we present our current knowledge on the cellular and molecular mechanisms governing the plasma membrane turnover rate of the transporter under resting conditions and in response to synaptic activity. We also discuss the notion that KCC2 lateral diffusion is one of the first parameters modulating the transporter membrane stability, allowing for rapid adaptation of Cl− transport to changes in neuronal activity. [ABSTRACT FROM AUTHOR]

Abstract: Altered brain metabolism is likely to be an important contributor to normal cognitive decline and brain pathology in elderly individuals. To characterize the metabolic changes associated with normal brain aging, we used high-field proton magnetic resonance spectroscopy in vivo to quantify 20 neurochemicals in the hippocampus and sensorimotor cortex of young adult and aged rats. We found significant differences in the neurochemical profile of the aged brain when compared with younger adults, including lower aspartate, ascorbate, glutamate, and macromolecules, and higher glucose, myo-inositol, N-acetylaspartylglutamate, total choline, and glutamine. These neurochemical biomarkers point to specific cellular mechanisms that are altered in brain aging, such as bioenergetics, oxidative stress, inflammation, cell membrane turnover, and endogenous neuroprotection. Proton magnetic resonance spectroscopy may be a valuable translational approach for studying mechanisms of brain aging and pathology, and for investigating treatments to preserve or enhance cognitive function in aging. [Copyright &y& Elsevier]

Directed cell migration and axonal guidance are essential steps in neural development that share many molecular mechanisms. The guidance of developing axons and migrating neurons is likely to depend on the precise control of plasmalemma turnover in selected regions of leading edges and growth cones, respectively. Previous results provided evidence of a signaling mechanism that couples chemotropic deleted in colorectal cancer (DCC)/Netrin-1 axonal guidance and exocytosis through Syntaxin1(Sytx1)/TI-VAMP SNARE proteins. Here we studied whether Netrin-1-dependent neuronal migration relies on a similar SNARE mechanism. We show that migrating neurons in the lower rhombic lip (LRL) express several SNARE proteins, and that DCC co-associates with Sytx1 and TI-VAMP in these cells. We also demonstrate that cleavage of Sytx1 by botulinum toxin C1 (BoNT/C1) abolishes Netrin-1-dependent chemoattraction of migrating neurons, and that interference of Sytx1 functions with shRNAs or Sytx1-dominant negatives disrupts Netrin-1-dependent chemoattraction of LRL neurons. These findings indicate that a Sytx1/DCC interaction is required for Netrin-1 guidance of migrating neurons, thereby highlighting a relationship between guidance signaling and SNARE proteins that regulate membrane turnover. [ABSTRACT FROM AUTHOR]

Photoreceptor membrane breakdown at dawn in the posterior median eyes of the spider Dinopis is described. Coated and smooth vesicles are shed into the receptor cytoplasm and are assembled into multivesicular bodies of two kinds: (i) Coated vesicles form loosely-assembled multivesicular bodies (mvbs) whose bounding membranes are derived from endoplasmic reticulum. (ii) Smooth vesicles generated by the mass disintegration of membrane aggregate to yield tightly-assembled multivesicular bodies which are not membrane-bound. Both types are either lysed in the inter-rhabdomeral cytoplasm, or degrade via multi-lamellar bodies to residual bodies (rbs) while they are being transported to the intermediate segments. Two systems are associated with lysis. ' Nebenkerne' produced by the rapid differentiation of GERL in the intermediate segments fuse with membrane-bound mvbs or rbs and may inject them with hydrolases. Partially-differentiated rigid tubules (Blest et al., 1978) travel to the receptive segments together with RER from the intermediate segments and also fuse with or engulf mvbs. Both systems may contain pro-enzymes which are activated at their target sites. No evidence of a close or necessary geometrical relationship between GERL and Golgi bodies has been seen, and there is no clear demarcation between RER, smooth ER and GERL which is entering into continuity with or engulfing mvbs. The implications of these findings for hypotheses about the origins of isolation membranes and autolytic systems in invertebrate systems are briefly discussed. [ABSTRACT FROM AUTHOR]

The fine structure of the principal compound eye of the shrimp, Palaemonetes, was studied under conditions of light and dark adaptation. Ommatidium structure and the response of pigmented cells to dark adaptation are similar to the situation in other decapod crustaceans. Light and dark adapted eyes differ in that the rhabdom changes its shape; morphological evidence suggests a possible sequence of events involving production, utilization, and degradation of photoreceptor membrane, a discontinuous process occurring only during changes from light to dark and dark to light. A hypothesis of membrane turnover is proposed. [ABSTRACT FROM AUTHOR]

A spontaneous mutation arising in Festuca pratensis has the effect of stabilizing the pigmentproteolipid complexes of thylakoid membranes so that leaf tissue does not turn yellow during senescence. Inheritance of the non-yellowing character was analysed in crosses between the wild-type cultivar Rossa and a mutant line Bf 993. Electrophoretic variants of cytoplasmic phosphoglucoisomerase coded by alleles of the nuclear gene Pgi-2 were used to identify hybrids during intercrossing. About 96% of the F progeny were heterozygous and all were phenotypically yellowing. In the F generation yellow ∶ green segregated in a ratio of 2.14∶1, not significantly different from 3∶1. In the backcross between F and Bf 993 the ratio was 1∶1 yellow ∶ green. There was no indication of linkage to Pgi-2. Senescence of detached Bf 993 and Rossa leaves was compared with that of the F hybrid. The hybrid behaved in an essentially identical fashion to the wildtype parent, and in marked contrast to the mutant, in all aspects of the senescence syndrome investigated, including loss of chlorophyll, carotenoids and the light-harvesting chlorophyll-protein of thylakoid membranes, and elevation of the particulate protein ∶ chlorophyll ratio in the terminal stages. It is concluded that there exists in Festuca pratensis a nuclear gene, designated Sid (senescence-induced degradation) which regulates turnover of hydrophobic components of photosynthetic membranes in ageing leaf tissue and which occurs in at least two allelic forms, y (yellow) dominant over g (green). [ABSTRACT FROM AUTHOR]

The maintenance of photoreceptor cell membranes in the blowfly was investigated in relation to the diurnal cycle, age, and the rpa (receptor potential absent) phototransduction mutation. The effect of disturbed membrane assembly on the electrical membrane properties was examined using single-electrode discontinuous current-clamp techniques. In wild-type flies the cross-sectional dimensions of the rhabdomeres were markedly reduced with age, and the quantity of synthetic organelles decreased concurrently, whereas no correlation was found between the diurnal cycle and membrane turnover. The rpa mutation is thought to block the visual transduction cascade in photoreceptor cells and to lead to degeneration of the photoreceptor cell bodies. The volume of rhabdomeres decreased markedly in rpa mutants and the quantity of synthetic organelles was reduced significantly, indicating an imbalance between photoreceptive membrane renewal and degradation. Also, the plasma membrane underwent degenerative changes. The passive electrical properties of photoreceptor cells - resting membrane voltages and input resistances - were only slightly changed from those of wild-type flies, although the photoreceptive membrane did not depolarize in response to light. This indicates no apparent disturbance in the function of the ionic channels in these membranes. Taken together, these results suggest that the photoreceptor cells need a functional phototransduction cascade with its feedback controls to maintain continuous renewal of rhabdomeres, but that the plasma membrane maintains its normal electrochemical properties despite extreme morphological degeneration of photoreceptor cell. [ABSTRACT FROM AUTHOR]

In order to study the synthesis and degradation processes of the photoreceptor membranes in the abalone, Nordotis discus, the localization of acyltransferase and acid hydrolase activities, respectively, were determined at the electron-microscopic level. Acyltransferase activity was localized on the cytoplasmic sides of thick (>10 nm) membranes of the following organelles: a few cisternae at the trans (or concave) side of Golgi apparatus, Golgi and probably related vesicles, short tubules, curved pentalaminar disks and limiting membranes of the phagosomal multivesicular bodies; all organelles were scattered in the peri- to supranuclear cytoplasm. The phospholipids, which are major components of the photoreceptor membrane, are considered to be synthesized by these membranes. Acid phosphatase activity was localized in the lumina of Golgi cisternae and vesicles, lysosomes, and smaller multivesicular and related bodies, but not in multilamellar bodies. The matrices of the larger multivesicular bodies and of the pigment granule complexes showed arylsulfatase activity. Vesiculated and autophagocytosed photoreceptor microvilli seemed to be degraded by acid hydrolases, forming multivesicular and related bodies. Supporting cells also showed acyltransferase and acid hydrolase activities. [ABSTRACT FROM AUTHOR]

Many crabs possess fused rhabdoms which are partly broken down at dawn and re-synthesised at dusk. The cross-sectional area of the rhabdom is therefore smaller during the day than at night. The only previously described mechanism of membrane removal from the rhabdomere in Crustacea involves the formation of pinocytotic vesicles at the bases of the microvilli. The geometry of the rhabdom is such that uniform pinocytosis across the base of each rhabdomere would result in a stack of orthogonally oriented rectangles. In the process described here, microvilli from the outer edges of the rhabdomeres are engulfed by adjacent retinula cells, reducing the number as well as the length of the microvilli and maintaining the smooth longitudinal profile needed for optimal functioning of the rhabdom. [ABSTRACT FROM AUTHOR]

The cross-sectional area of rhabdomeres in the compound eye of the blowfly, Lucilia, was found to remain constant under 12 h light/12 h dark cyclic lighting, and 10 days constant light or darkness. It was reduced only slightly during 3 h light after 10 days darkness (by 21%), or on exposure to 2h darkness + 1.5 h light after 10 days light (by 10%). Morphological evidence indicates that shedding of photoreceptor membrane during turnover is achieved by an extracellular route, and by pinocytosis from the bases of the microvilli. The photoreceptor membrane shed by both mechanisms appears to accumulate in multivesicular bodies. The amount of photoreceptor membrane shedding, as indicated by numbers of multivesicular bodies, is constant throughout the day and night on cyclic lighting, decreases in constant darkness, but returns to its normal level after an exposure to 3 h light subsequent to 10 days darkness. [ABSTRACT FROM AUTHOR]

In the compound eye of the locust, Locusta, the cross-sectional area of the rhabdoms increases at 'dusk' by 4.7-fold due to the rapid assembly of new microvillar membrane, and decreases at 'dawn' by a corresponding amount as a result of pinocytotic shedding from the microvilli. The rhabdoms at night have more and longer photoreceptor microvilli than rhabdoms during the day. The orientations of the six rhabdomeres that comprise the distal rhabdom also change. The density of intramembrane particles on the P-face of the microvillar membrane, putatively representing mostly rhodopsin molecules, or aggregates thereof, does not change. An alteration in the size of the ommatidial field-stop, produced by the primary pigment cells, is concomitant with the change in rhabdom size. At night the increase in size of the field-stop must widen the angular acceptance of a rhabdom, increasing the capture of photons from an extended field. Conversely, during the day, when photons are more abundant, its decrease must narrow the acceptance angle, increasing angular resolution. Because of the presence of this field-stop, the optics of the ommatidium would not be greatly affected if the rhabdom were to remain always at its night size. It is argued, therefore, that the variable-size rhabdom must have resulted from some demand other than that of light/dark adaptation. Changes in size and organisation of the rhabdoms in response to various light regimes indicate that: (1) Rapid shedding of photoreceptor membrane is induced by the onset of light, but shedding also occurs slowly in darkness during the day. (2) Microvillar assembly is initiated by the onset of darkness, but also occurs at the normal time of dusk without a change in ambient lighting, provided there has been some light during the day. Therefore, both shedding and assembly of microvillar membrane are affected by the state of illumination, but also appear to be under some endogenous control. [ABSTRACT FROM AUTHOR]

In order to examine a diurnal (24 h) rhythm in the fine structure of photoreceptors of abalone, Nordotis discus, the animals were kept on a daily light-dark cycle (light period: 6:00 a.m.-6:00 p.m., 2-5 lx) for 10 days; subsequently their eyes were fixed at nine specified times of the day. At 5:30 a.m. the photoreceptors consisted of numerous terminal microvilli growing on the surface of a dome-shaped apical cytoplasmic process (AP). Illumination induced breakdown of the photoreceptive microvilli into many vesicles. These vesicles were densely aggregated on almost the entire surface of enlarged APs at 7:00 a.m. After 1 h they markedly decreased in number. The cell at 10:00 a.m. possessed sparse, short and irregularly-contoured microvilli. The microvilli gradually increased in number from the middle through the later half of the light period. Only within 1 h after deprivation of light shorter but characteristic terminal microvilli were observed. By midnight they became similar in fine structure to those at 5:30 a.m. [ABSTRACT FROM AUTHOR]

Large areas of photoreceptor membrane are synthesized in the retinula cells of the crab Leptograpsus variegatus at dusk. Initially, new membrane differentiates from rough endoplasmic reticulum (ER) as large tubules of smooth ER. These tubules transform to concentric ellipsoids of closely apposed pairs of membranes ('doublet ER'), sometimes passing through an intervening crenate form. The new membrane is transported through bridges of cytoplasm that cross the palisade to the rhabdom region, from which the remains of the rhabdomeres that were built during the previous dusk have been dissolved. The degradation of the old microvilli of one rhabdomere is accomplished without affecting neighbouring rhabdomeres of other cells. New microvilli are assembled in situ from sheets of doublet ER, which are converted to tubules oriented in the same direction as the future microvilli. The cytoplasmic face of the ER remains the cytoplasmic face of the tubules, which become progressively narrower, partly by further longitudinal division, until the final diameter of the microvillus is reached. A central core is often seen in transverse sections of mature microvilli. It may be involved in the final consolidation, but rhabdomeric microvilli are not formed in the same manner as those of intestinal brush border cells. There is no evidence that new membrane passes through the Golgi compartment before incorporation into the rhabdom, as is the case for rod outer segment membrane in vertebrate photoreceptors. [ABSTRACT FROM AUTHOR]

The effect of illumination on the degradation of microvillar membrane in the invertebrate photoreceptor cell has been correlated with the appearance in the cytoplasm of certain distinct lysosome-related bodies. Three types of organelles were distinguished in the retinula cell cytoplasm of the crayfish, multivesicular bodies (MVB), both large (4.20-1.50 μm) and small (1.49-0.30 μm), combination bodies (CB), and lamellar bodies (LB). Under diurnal lighting conditions significant temporal differences were found in the appearance of these three classes of organelles in the retinula cell. Small MVB are present at a consistent level throughout most of the diurnal cycle but show peak numbers at 30 min after light onset and again after 6 h of dark adaptation. Large MVB increase significantly 1 h after light onset and remain elevated through 4 h in the light. After 4 h the large MVB decline gradually for the remaining light period. Combination bodies and LB do not begin to increase until 1 h after light onset and are at peak levels between 4 and 6 h into the light period. The minimum rhabdome diameter coincides with the peak levels of large MVB, CB, and LB. These data support the hypothesis that light causes microvillar membrane breakdown, resulting in the initial production of MVB which in turn undergo degradation to form CB and finally LB. This primary degradative response appears to be completed within the first 8 h of the light period. [ABSTRACT FROM AUTHOR]

To investigate the turnover of complete junctional membrane areas ('domain turnover'), psoriatic epidermis, keratoacanthoma, and squamous cell carcinoma were studied with an electron microscope and compared with the normal epidermis. Three types of domain turnover of junctions were observed. First, the uptake of intact intercellular junctions, which occurred as complete or incomplete annular junctions in the cytoplasm. Mainly gap junctions were involved. Second, the formation of autojunctions bridging an infolding at the surface of a cell, which is then incorporated and occurs partly with an attached saccule in the cytoplasm. This type was confined to desmosomes. Finally, the loss of intercellular desmosomes into the intercellular space was observed. Proliferating epidermis was associated with an increased domain turnover of gap junctional membrane areas. Domain turnover of desmosomes prevailed in tumors. [ABSTRACT FROM AUTHOR]

The diurnal changes of rhabdom structures under a regular light cycle (L:D=12:12) have been examined in the eyes of the grapsid carb, . Night rhabdoms are larger in diameter than noontime rhabdoms by more than a factor of 2. The rhabdomeric membranes are internalized around dawn as coated vesicles. The coated vesicles may be transferred to the parent cytoplasm following transformation into sickle-shaped vesicles or double membrane-bounded vesicles. Conversely, the increase of rhabdom size from noon to midnight may be largely due to the assembly of vacuoles into rhabdomeric membranes. In addition to the diurnal change in rhabdom size, adjacent rhabdomerlc microvilli appear to easily fuse to each other to form thick microvilli, which re-divide again into normal ones. Moreover, extracellular shedding of microvillar membranes occurs during the dark period. Observed changes of rhabdomeric microvilli are discussed in relation to turnover of the photosensitive membranes. [ABSTRACT FROM PUBLISHER]

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Capacitance changes in cell-attached patches of rat melanotrophs were measured by a high-frequency lock-in amplifier. The background noise of around 30 aF allowed the detection of discrete steps due to fission (endocytosis) and fusion (exocytosis) of vesicles with diameters as small as 60 nm. The amplitude of both types of steps was similar with a modal value of around 300 aF. The frequency of these steps was not changed, if secretagougues such as ionomycin and/or dibutyril cAMP, were applied to the bathing solution. Moreover, this treatment did not result in an increased appearance of expected 2000 to 3000 aF steps due to exocytosis of secretory granules. We conclude that the likely explanation for recorded capacitance steps is that they represent the constitutive vesicle traffic. From the typical frequency and amplitude of these events (around 1 min–1, 300 aF) in a membrane patch (26 fF) it is estimated that the whole membrane of a rat melanotroph may be ingested under our conditions in 1 to 2 h. [ABSTRACT FROM AUTHOR]

Light stimulation of locust photoreceptors causes a translocation of submicrovillar cisternae of the endoplasmic reticulum away from the rhabdome, and a movement of mitochondria towards the rhabdome. To examine whether the microtubule cytoskeleton could be involved in these organelle movements, we have analysed the distribution of the microtubule-dependent motor proteins kinesin and cytoplasmic dynein in the retina of the locust Schistocerca gregaria. Both kinesin and cytoplasmic dynein are associated with vesicular structures that are distinct from mitochondria and the submicrovillar endoplasmic reticulum. These results, together with the previous demonstration of a lack of microtubules in the cell area of light-dependent organelle movements, provide evidence that the microtubule cytoskeleton is not involved with light-induced organelle translocations. [ABSTRACT FROM AUTHOR]

Phospholipase B-mediated hydrolysis of phosphatidylcholine (PC) results in the formation of free fatty acids and glycerophosphocholine (GPC) in the yeast Saccharomyces cerevisiae GPC can be reacylated by the glycerophosphocholine acyltransferase Gpc1, which produces lysophosphatidylcholine (LPC), and LPC can be converted to PC by the lysophospholipid acyltransferase Ale1. Here, we further characterized the regulation and function of this distinct PC deacylation/reacylation pathway in yeast. Through in vitro and in vivo experiments, we show that Gpc1 and Ale1 are the major cellular GPC and LPC acyltransferases, respectively. Importantly, we report that Gpc1 activity affects the PC species profile. Loss of Gpc1 decreased the levels of monounsaturated PC species and increased those of diunsaturated PC species, whereas Gpc1 overexpression had the opposite effects. Of note, Gpc1 loss did not significantly affect phosphatidylethanolamine, phosphatidylinositol, and phosphatidylserine profiles. Our results indicate that Gpc1 is involved in postsynthetic PC remodeling that produces more saturated PC species. qRT-PCR analyses revealed that GPC1 mRNA abundance is regulated coordinately with PC biosynthetic pathways. Inositol availability, which regulates several phospholipid biosynthetic genes, down-regulated GPC1 expression at the mRNA and protein levels and, as expected, decreased levels of monounsaturated PC species. Finally, loss of GPC1 decreased stationary phase viability in inositol-free medium. These results indicate that Gpc1 is part of a postsynthetic PC deacylation/reacylation remodeling pathway (PC-DRP) that alters the PC species profile, is regulated in coordination with other major lipid biosynthetic pathways, and affects yeast growth., (© 2019 Anaokar et al.)

SENSITIVE TO FREEZING 2 (SFR2) is classified as a family I glycosyl hydrolase but has recently been shown to have galactosyltransferase activity in Arabidopsis thaliana. Natural occurrences of apparent glycosyl hydrolases acting as transferases are interesting from a biocatalysis standpoint, and knowledge about the interconversion can assist in engineering SFR2 in crop plants to resist freezing. To understand how SFR2 evolved into a transferase, the relationship between its structure and function are investigated by activity assay, molecular modeling, and site-directed mutagenesis. SFR2 has no detectable hydrolase activity, although its catalytic site is highly conserved with that of family 1 glycosyl hydrolases. Three regions disparate from glycosyl hydrolases are identified as required for transferase activity as follows: a loop insertion, the C-terminal peptide, and a hydrophobic patch adjacent to the catalytic site. Rationales for the effects of these regions on the SFR2 mechanism are discussed., (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)