Phase and amplitude reconstruction in single-pixel transmission microscopy: a comparison of Hadamard, cosine, and noiselet bases.

Hadamard, cosine, and noiselet bases are implemented into a digital holographic microscope based on single-pixel imaging with the capability to retrieve images of complex objects. The object is illuminated with coherent light modulated with different patterns deployed in a digital micromirror device, and the resulting fields are captured by single-pixel detection. For amplitude images, the experimental results of the three bases are evaluated with the peak SNR criteria. It is shown that the cosine basis recovers amplitude distributions with the best quality. Regarding phase images, the recovered ones compare well with those obtained with a CMOS camera.

Real-time reflectance anisotropy spectroscopy of GaAs homoepitaxy.

Reflectance anisotropy spectroscopy (RAS) is a highly sensitive optical probe for the real-time study of the epitaxial growth of zincblende semiconductors. Here we report on (1) non-equilibrium RAS spectra acquired in real time during the homoepitaxial growth of GaAs, and (2) RAS spectra for GaAs surfaces under equilibrium with several arsenic overpressures. We show that in both cases RAS spectra can be decomposed into two basic components, each with a characteristic line shape. We further show that both dynamic and equilibrium RAS spectra are described by the same pair of basic components. We conclude that the time evolution of non-equilibrium RAS spectra acquired during the epitaxial growth can be described in terms of RAS spectra for equilibrium surfaces. The results reported here should be useful for the interpretation of the physics underlying the rapid time evolution of dynamic RAS spectra during the first monolayer growth. Thus, we show that RAS constitutes a valuable tool for the study of epitaxial growth mechanisms.

Solar wind thermally induced magnetic fluctuations.

A kinetic description of Alfvén-cyclotron magnetic fluctuations for anisotropic electron-proton quasistable plasmas is studied. An analytical treatment, based on the fluctuation-dissipation theorem, consistently shows that spontaneous fluctuations in plasmas with stable distributions significantly contribute to the observed magnetic fluctuations in the solar wind, as seen, for example, in [S. D. Bale et al., Phys. Rev. Lett. 103, 211101 (2009)], even far below from the instability thresholds. Furthermore, these results, which do not require any adjustable parameters or wave excitations, are consistent with the results provided by hybrid simulations. It is expected that this analysis contributes to our understanding of the nature of magnetic fluctuations in the solar wind.

Two predicted α-helices within the prion-like domain of TIAR-1 play a crucial role in its association with stress granules in Caenorhabditis elegans.

Stress granules (SGs) are sites for mRNA storage, protection, and translation repression. TIA1 and TIAR1 are two RNA-binding proteins that are key players in SGs formation in mammals. TIA1/TIAR have a prion-like domain (PrD) in their C-terminal that promotes liquid-phase separation. Lack of any TIA1/TIAR has severe consequences in mice. However, it is not clear whether the failure to form proper SGs is the cause of any of these problems. We disrupted two predicted α-helices within the prion-like domain of the Caenohabditis elegans TIA1/TIAR homolog, TIAR-1, to test whether its association with SGs is important for the nematode. We found that tiar-1 PrD mutant animals continued to form TIAR-1 condensates under stress in the C. elegans gonad. Nonetheless, TIAR-1 condensates appeared fragile and disassembled quickly after stress. Apparently, the SGs continued to associate regularly as observed with CGH-1, an SG marker. Like tiar-1-knockout nematodes, tiar-1 PrD mutant animals exhibited fertility problems and a shorter lifespan. Notwithstanding this, tiar-1 PrD mutant nematodes were no sensitive to stress. Our data demonstrate that the predicted prion-like domain of TIAR-1 is important for its association with stress granules. Moreover, this domain may also play a significant role in various TIAR-1 functions unrelated to stress, such as fertility, embryogenesis and lifespan.

Data reduction for spatially resolved reflectance anisotropy spectrometer.

We show that in spatially resolved reflectance anisotropy (RA) spectrometers, off-axis optical rays introduce a spurious signal component that cannot be addressed by optical alignment. Such a component is associated with the difference between the reflectivities s and p of the sample and depends, in a complex manner, on the incidence position of the incident light on the surface of the sample. We report a data-reduction procedure to easily identify and remove spurious RA signals associated with the off-axis optical rays, based on the singular value decomposition analysis of spatially resolved RA spectra. We validated this approach by developing a spatially resolved RA spectrometer based on an 8 × 8 multi-anode photomultiplier (PMT). The PMT allowed the use of phase-sensitive detection techniques to enhance the signal-to-noise ratio, which is essential for the evaluation of the proposed data reduction procedure.

Microreflectance difference spectrometer based on a charge coupled device camera: surface distribution of polishing-related linear defect density in GaAs (001).

We describe a microreflectance difference (microRD) spectrometer based on a charge coupled device (CCD), in contrast to most common RD spectrometers that are based on a photomultiplier or a photodiode as a light detector. The advantage of our instrument over others is the possibility to isolate the RD spectrum of specific areas of the sample; thus topographic maps of the surface can be obtained. In our setup we have a maximum spatial resolution of approximately 2.50 microm x 2.50 microm and a spectral range from 1.2 to 5.5 eV. To illustrate the performance of the spectrometer, we have measured strains in mechanically polished GaAs (001) single crystals.

Apoptosis contributes to protect germ cells from the oogenic germline starvation response but is not essential for the gonad shrinking or recovery observed during adult reproductive diapause in C. elegans.

When C. elegans hermaphrodites are deprived of food during the mid-L4 larval stage and throughout adulthood, they enter an alternative stage termed "adult reproductive diapause (ARD)" in which they halt reproduction and extend their lifespan. During ARD, germ cell proliferation stops; oogenesis is slowed; and the gonad shrinks progressively, which has been described as the "oogenic germline starvation response". Upon refeeding, the shrunken gonad is regenerated, and animals recover fertility and live out their remaining lifespan. Little is known about the effects of ARD on oocyte quality after ARD. Thus, the aim of this study was to determine how oocyte quality is affected after ARD by measuring brood size and embryonic lethality as a reflection of defective oocyte production. We found that ARD affects reproductive capacity. The oogenic germline starvation response protects oogenic germ cells by slowing oogenesis to prevent prolonged arrest in diakinesis. In contrast to a previous report, we found that germ cell apoptosis is not the cause of gonad shrinkage; instead, we propose that ovulation contributes to gonad shrinkage during the oogenic germline starvation response. We show that germ cell apoptosis increases and continues during ARD via lin-35/Rb and an unknown mechanism. Although apoptosis contributes to maintain germ cell quality during ARD, we demonstrated that apoptosis is not essential to preserve animal fertility. Finally, we show that IIS signaling inactivation partially participates in the oogenic germline starvation response.

Stress-induced germ cell apoptosis by a p53 independent pathway in Caenorhabditis elegans.

In Caenorhabditis elegans, several distinct apoptosis pathways have been characterized in the germline. The physiological pathway is though to eliminate excess germ cells during oogenesis to maintain gonad homeostasis and it is activated by unknown mechanisms. The DNA damage-induced germ cell apoptosis occurs in response to genotoxic agents and involves the proteins EGL-1 and CED-13, and the DNA damage response protein p53. Germ cell apoptosis can also be induced in response to pathogen infection through an EGL-1 dependent pathway. To gain insight into the mechanism and functions of germ cell apoptosis, we investigated whether and how other forms of stress induce this cell death. We found that oxidative, osmotic, heat shock and starvation stresses induce germ cell apoptosis through a p53 and EGL-1 independent pathway. We also learned that the MAPK kinases MEK-1 and SEK-1, and the p53 antagonist protein ABL-1, are essential for stress-induced germ cell apoptosis. We conclude that in C. elegans responses to various stresses that do not involve genotoxicity include an increase in germ cell apoptosis through the physiological pathway.

Caenorhabditis elegans ATPase inhibitor factor 1 (IF1) MAI-2 preserves the mitochondrial membrane potential (Δψm) and is important to induce germ cell apoptosis.

When the electrochemical proton gradient is disrupted in the mitochondria, IF1 (Inhibitor Factor-1) inhibits the reverse hydrolytic activity of the F1Fo-ATP synthase, thereby allowing cells to conserve ATP at the expense of losing the mitochondrial membrane potential (Δψm). The function of IF1 has been studied mainly in different cell lines, but these studies have generated contrasting results, which have not been helpful to understand the real role of this protein in a whole organism. In this work, we studied IF1 function in Caenorhabditis elegans to understand IF1´s role in vivo. C. elegans has two inhibitor proteins of the F1Fo-ATPase, MAI-1 and MAI-2. To determine their protein localization in C. elegans, we generated translational reporters and found that MAI-2 is expressed ubiquitously in the mitochondria; conversely, MAI-1 was found in the cytoplasm and nuclei of certain tissues. By CRISPR/Cas9 genome editing, we generated mai-2 mutant alleles. Here, we showed that mai-2 mutant animals have normal progeny, embryonic development and lifespan. Contrasting with the results previously obtained in cell lines, we found no evident defects in the mitochondrial network, dimer/monomer ATP synthase ratio, ATP concentration or respiration. Our results suggest that some of the roles previously attributed to IF1 in cell lines could not reflect the function of this protein in a whole organism and could be attributed to specific cell lines or methods used to silence, knockout or overexpress this protein. However, we did observe that animals lacking IF1 had an enhanced Δψm and lower physiological germ cell apoptosis. Importantly, we found that mai-2 mutant animals must be under stress to observe the role of IF1. Accordingly, we observed that mai-2 mutant animals were more sensitive to heat shock, oxidative stress and electron transport chain blockade. Furthermore, we observed that IF1 is important to induce germ cell apoptosis under certain types of stress. Here, we propose that MAI-2 might play a role in apoptosis by regulating Δψm. Additionally, we suggest that IF1 function is mainly observed under stress and that, under physiological conditions, this protein does not play an essential role.

Note: A simple multi-channel optical system for modulation spectroscopies.

Photoreflectance-difference (PR/PRD) and reflectance-difference (RD) spectroscopies employ synchronic detection usually with lock-in amplifiers operating at moderate (200-1000 Hz) and high (50-100 KHz) modulation frequencies, respectively. Here, we report a measurement system for these spectroscopies based on a multichannel CCD spectrometer without a lock-in amplifier. In the proposed scheme, a typical PRD or RD spectrum consists of numerical subtractions between a thousand CCD captures recorded, while a photoelastic modulator is either operating or inhibited. This is advantageous and fits the slow response of CCD detectors to high modulation frequencies. The resulting spectra are processed with Savitzky-Golay filtering and compared well with those measured with conventional scanning systems based on lock-in amplifiers.

On a growth model for complex networks capable of producing power-law out-degree distributions with wide range exponents.

The out-degree distribution is one of the most reported topological properties to characterize real complex networks. This property describes the probability that a node in the network has a particular number of outgoing links. It has been found that in many real complex networks the out-degree has a behavior similar to a power-law distribution, therefore some network growth models have been proposed to approximate this behavior. This paper introduces a new growth model that allows to produce out-degree distributions that decay as a power-law with an exponent in the range from 1 to ∞.

LIN-35/Rb causes starvation-induced germ cell apoptosis via CED-9/Bcl2 downregulation in Caenorhabditis elegans.

Apoptosis is an important mechanism for maintaining germ line health. In Caenorhabditis elegans, germ cell apoptosis occurs under normal conditions to sustain gonad homeostasis and oocyte quality. Under stress, germ cell apoptosis can be triggered via different pathways, including the following: (i) the CEP-1/p53 pathway, which induces germ cell apoptosis when animals are exposed to DNA damage; (ii) the mitogen-activated protein kinase kinase (MAPKK) pathway, which triggers germ cell apoptosis when animals are exposed to heat shock, oxidative stress, or osmotic stress; and (iii) an unknown mechanism that triggers germ cell apoptosis during starvation. Here, we address how starvation induces germ cell apoptosis. Using polysomal profiling, we found that starvation for 6 h reduces the translationally active ribosomes, which differentially affect the mRNAs of the core apoptotic machinery and some of its regulators. During starvation, lin-35/Rb mRNA increases its expression, resulting in the accumulation of this protein. As a consequence, LIN-35 downregulates the expression of the antiapoptotic gene ced-9/Bcl-2. We observed that the reduced translation of ced-9/Bcl-2 mRNA during food deprivation together with its downregulation drastically affects its protein accumulation. We propose that CED-9/Bcl-2 downregulation via LIN-35/Rb triggers germ cell apoptosis in C. elegans in response to starvation.

A rapid reflectance-difference spectrometer for real-time semiconductor growth monitoring with sub-second time resolution.

We report on a rapid, 32-channel reflectance-difference (RD) spectrometer with sub-second spectra acquisition times and ΔR/R sensitivity in the upper 10(-4) range. The spectrometer is based on a 50 kHz photo-elastic modulator for light polarization modulation and on a lock-in amplifier for signal harmonic analysis. Multichannel operation is allowed by multiplexing the 32 outputs of the spectrometer into the input of the lock-in amplifier. The spectrometer spans a wavelength range of 230 nm that can be tuned to cover E(1) and E(1) + Δ(1) transitions for a number of III-V semiconductors at epitaxial growth temperatures, including GaAs, InAs, AlAs, and their alloys. We present two examples of real-time measurements to demonstrate the performance of the RD spectrometer, namely, the evolution of the RD spectrum of GaAs (001) annealed at 500 °C and the time-dependent RD spectrum during the first stages of the epitaxial growth of In(0.3)Ga(0.7)As on GaAs (001) substrates.

Strain oscillations probed with light.

We show that reflectance difference spectroscopy (RDS) is sensitive to the inhomogeneous surface and thin film strain which builds up during hetero- and homoepitaxial growth. The RDS signal is affected by the local, mean square atomic displacements in the substrate resulting from the stress relaxation of strained adlayer islands. For layer-by-layer growth an oscillatory variation of the RDS intensity is observed. These results demonstrate the potentiality of RDS to probe the growth kinetics on structurally anisotropic surfaces.

Measurement of the surface strain induced by reconstructed surfaces of GaAs (001) using photoreflectance and reflectance-difference spectroscopies.

We report photoreflectance-difference and reflectance-difference measurements on reconstructed GaAs (001) surfaces. From these data the linear and quadratic electro-optic coefficient spectra are determined in the important 2.8-3.4 eV spectral region. The surface strain and fields induced by the surface reconstruction are also determined. We show experimentally that between c(4x4) and (2x4) surfaces, there is an inversion of the surface electric field which we attribute to a direct piezo-electric effect related to the surface strain induced by reconstruction.

Posttranscriptional control mediates cell type-specific localization of catalase A during Aspergillus nidulans development.

Two differentially regulated catalase genes have been identified in the fungus Aspergillus nidulans. The catA gene belongs to a class whose transcripts are specifically induced during asexual sporulation (conidiation) and encodes a catalase accumulated in conidia. Using a developmental mutant affected in the brlA gene, which is unable to form conidia but capable of producing sexual spores (ascospores), we demonstrated that the catA mRNA accumulated during induction of conidiation but did not produce CatA protein. In contrast, high levels of catalase A activity were detected in the ascospores produced by this mutant, indicating that the catA gene is posttranscriptionally regulated. The same type of regulation was observed for a catA::lacZ translational gene fusion, suggesting that the catA message 5' untranslated region could be involved in translational control during development. In a wild-type strain, beta-galactosidase activity driven from the catA::lacZ gene fusion was low in hyphae and increased 50-fold during conidiation and 620-fold in isolated conidia. Consistent with this finding spatial expression of the reporter gene was restricted to metulae, phialides, and conidia. Conidium-associated expression was maintained in a stuA mutant, in which the conidiophore cell pattern is severely deranged. catA mRNA accumulation was also observed when vegetative mycelia was subject to oxidative, osmotic, and nitrogen or carbon starvation stress. Nevertheless, catalase A activity was restricted to the conidia produced under nutrient starvation. Our results provide support for a model in which translation of the catA message, accumulated during conidiation or in response to different types of stress, is linked to the morphogenetic processes involved in asexual and sexual spore formation. Our findings also indicate that brlA-independent mechanisms regulate the expression of genes encoding spore-specific products.

Increased transformation frequency and tagging of developmental genes in Aspergillus nidulans by restriction enzyme-mediated integration (REMI).

We have used a plasmid containing the argB gene to transform an Aspergillus nidulans argB-deleted strain in the presence of restriction enzymes and show a 20- to 60-fold increase in transformation frequency via restriction enzyme-mediated integration (REMI). This procedure was used to try to tag new genes involved in the asexual development of this fungus. More than 2000 transformants isolated following electroporation of conidia and approximately 3700 transformants recovered following protoplast fusion were screened for sporulation defects. Unexpectedly, developmental mutants were obtained only when the protoplast fusion approach was used. Southern blot analysis of these mutants, and of randomly selected transformants obtained by electroporation, was consistent with the occurrence of single plasmid integration events in 33 and 65% of the cases, respectively. The argB marker was shown to be tightly linked to the mutant phenotype in only 62% of the mutants analyzed by sexual crosses. Partial DNA sequencing of a tagged gene, whose mutation delays asexual sporulation and results in a fluffy phenotype, showed no homology to previously reported sequences. Our results indicate that REMI can be used in A. nidulans to increase the transformation frequency and illustrate the advantages and potential problems when using REMI to tag genes of interest in this and other fungi.

catA, a new Aspergillus nidulans gene encoding a developmentally regulated catalase.

Aspergillus nidulans asexual sporulation (conidiation) is a model system for studying gene regulation and development. The CAN5 cDNA is one of several clones isolated based on transcript induction during conidiation. Here we present the molecular characterization of its corresponding gene, demonstrating that it encodes a developmentally regulated catalase, designated catA. The catA 744-amino-acid-residue polypeptide shows significant identity to other catalases. Its similarity to prokaryotic catalases is greater than to other fungal catalases. catA mRNA is barely detectable in growing mycelia, highly induced during sporulation, and present in isolated spores. However, catA expression is not dependent on the developmental regulatory genes brlA, abaA and wetA. Direct catalase activity determination in native gels revealed the existence of two bands of activity. One of these bands represented the major activity during vegetative growth and was induced during sporulation. The second catalase activity appeared after the induction of sporulation and was the predominant activity in spores. Disruption of catA abolished the major spore catalase without eliminating the vegetative activity, indicating the existence of at least two catalase genes in A. nidulans. catA-disrupted mutants produced spores that were sensitive to H2O2, as compared to wild-type spores. The increase in the activity of the vegetative catalase and the appearance of a second catalase during asexual sporulation is consistent with the occurrence of an oxidative stress during development.

cgh-1, a conserved predicted RNA helicase required for gametogenesis and protection from physiological germline apoptosis in C. elegans.

A high frequency of apoptosis is a conserved hallmark of oocyte development. In C. elegans, about half of all developing oocytes are normally killed by a physiological germline-specific apoptosis pathway, apparently so that they donate cytoplasm to the survivors. We have investigated the functions of CGH-1, the C. elegans ortholog of the predicted RNA helicase ste13/ME31B/RCK/p54, which is germline-associated in metazoans and required for sexual reproduction in yeast. We show that CGH-1 is expressed specifically in the germline and early embryo, and is localized to P granules and other possible mRNA-protein particles. cgh-1 is required for oocyte and sperm function. It is also needed to prevent the physiological germline apoptosis mechanism killing essentially all developing oocytes, making lack of cgh-1 function the first stimulus identified that can trigger this mechanism. We conclude that cgh-1 and its orthologs may perform conserved functions during gametogenesis, that in C. elegans certain aspects of oocyte development are monitored by the physiological germline apoptosis pathway, and that similar surveillance mechanisms may contribute to germline apoptosis in other species.

Enhanced optical sensitivity to adsorption due to depolarization of anisotropic surface states.

Reflectance difference spectroscopy is used to probe the optical transitions between surface states on the Cu(110) surface. Upon deposition of smallest amounts of carbon monoxide (CO) the signal is strongly quenched, which translates into a huge cross section of the order of 1000 A(2) for a single adsorbed CO molecule. This strongly enhanced surface sensitivity is interpreted as the loss in anisotropy (depolarization) of the surface states due to scattering from the adsorbed CO molecules. This feature renders RDS an extremely sensitive tool to probe the adsorption kinetics on anisotropic metal surfaces.