Urea Extraction Research Articles

Development of a functional in vitro integration system for an integral membrane protein, SecG

A functional in vitro integration system for an integral membrane protein, SecG, comprising an efficient translation system supplemented with inverted membrane vesicles (IMV) was developed. When SecG was synthesized in the presence of IMV prepared from a Δ secG strain (ΔSecG IMV), the synthesized SecG was recovered with the IMV. A population of SecG was resistant to urea extraction, indicating that the synthesized SecG was integrated into ΔSecG IMV. Addition of signal recognition particle and its receptor (SRP) and SecA caused an increase in the amount of the urea-resistant form of SecG. When IMV into which SecG had been integrated were subjected to the translocation assay, the translocation activity was found to be significantly stimulated compared with for ΔSecG IMV. Moreover, when SRP and SecA had been supplemented, the translocation activity nearly recovered to the level in IMV prepared from the wild type strain. These results indicate that the in vitro synthesized SecG could be functionally integrated into ΔSecG IMV with the help of SRP and SecA. We also present evidence that the membrane targeting and integration of SecG is stimulated by externally added SecA and SecG itself.

Optimization of medium constituents for improved chitinase production byPaenibacillussp. D1 using statistical approach

Statistical optimization of medium components for improved chitinase production by Paenibacillus sp. D1. Urea, K(2)HPO(4), chitin and yeast extract were identified as significant components influencing chitinase production by Paenibacillus sp. D1 using Plackett-Burman method. Response surface methodology (central composite design) was applied for further optimization. The concentrations of medium components for improved chitinase production were as follows (g l(-1)): urea, 0.33; K(2)HPO(4), 1.17; MgSO(4), 0.3; yeast extract, 0.65 and chitin, 3.75. This statistical optimization approach led to the production of 93.2 +/- 0.58 U ml(-1) of chitinase. The important factors controlling the production of chitinase by Paenibacillus sp. D1 were identified as urea, K(2)HPO(4), chitin and yeast extract. Statistical approach was found to be very effective in optimizing the medium components in manageable number of experimental runs with overall 2.56-fold increase in chitinase production. The present investigation provides a report on statistical optimization of medium components for improved chitinase production by Paenibacillus sp. D1. Paenibacillus species are gram-positive, spore-forming bacteria with several PGPR and biocontrol potentials. However, only few reports concerning mycolytic enzyme production especially chitinases are available. Chitinase produced by Paenibacillus sp. D1 represents new source for biotechnological and agricultural use.

Determination of an economical medium for growth ofLactobacillus fermentumusing response surface methodology

Lactobacillus fermentum is a widely utilized probiotic compound fed as an alternative to antibiotics for growth promotion in a wide variety of livestock species. The objective of this research is to develop an economical and practical fermentation medium for the growth of Lact. fermentum using response surface methodology. A two-level Plackett-Burman design was used to determine which factors in the fermentation medium influence the growth of Lact. fermentum. Under our experimental conditions, peptone, urea and yeast extract were found to be major factors. Then, the steepest ascent method and the central composite design were applied to optimize the culture of Lact. fermentum. The following composition of the fermentation medium was estimated to be the most economical formula (per litre): 30 g corn syrup, 15 g glucose, 14.4 g peptone, 7 g (NH(4))(2)SO(4), 0.5 g urea, 3 g sodium acetate, 4 g sodium citrate, 0.1 g MnSO(4).4H(2)O, 0.5 g MgSO(4).7H(2)O, 7.3 g yeast extract, 0.5 g K(2)HPO(4). Based on 10 side-by-side comparisons, we found that the yield of Lact. fermentum using our fermentation medium was 64% greater than those using modified de Man, Rogosa and Sharp broth (MRS) medium (1.8 x 10(9) CFU ml(-1)vs 1.1 x 10(9) CFU ml(-1), respectively), while the cost was 89% lower than MRS. This research indicates that it is possible to increase bacterial yield by using inexpensive materials. It is more likely that the use of Lact. fermentum as a probiotic will increase. The low cost medium developed in this research can be used for large-scale, commercial application where economics are quite likely to be important.

Interaction of different media on production and biocontrol efficacy of Pseudomonas fluorescens P-35 and Bacillus subtilis B-3 against grey mould of apple.

SUMMARY In this study, growth and efficacy of two biological control agents, Pseudomonas fluorescens P-35 and Bacillus subtilis B-3, were evaluated in combinations of two carbon (sucrose and molasses) and two nitrogen (urea and yeast extract) sources to optimize control of Botrytis cinerea on apple. Nutrient broth was used as control. Results indicated that the medium containing molasses and yeast extract (MY) in a 1:1 w/w ratio supported rapid growth and high cell yields in both strains. The biocontrol efficacy of P-35 grown in MY was higher than when grown in the other media, and the percentage of decayed fruits was 38.6% as compared to the control (100%), after ten days incubation. P-35 also showed good biocontrol efficacy in MY after 20 days. Nutrient broth resulted in poor biocontrol efficacy for B-3 after ten days, as compared with the other media. For both strains molasses were a good carbon source in combination with yeast extract, but antagonist growth was decreased when molasses were complemented with urea. The results obtained could provide a reliable basis for mass multiplication of biocontrol agents in fermentation processes.

Expression of TDP-43 C-terminal Fragments in Vitro Recapitulates Pathological Features of TDP-43 Proteinopathies

The disease protein in frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U) and amyotrophic lateral sclerosis (ALS) was identified recently as the TDP-43 (TAR DNA-binding protein 43), thereby providing a molecular link between these two disorders. In FTLD-U and ALS, TDP-43 is redistributed from its normal nuclear localization to form cytoplasmic insoluble aggregates. Moreover, pathological TDP-43 is abnormally ubiquitinated, hyperphosphorylated, and N-terminally cleaved to generate C-terminal fragments (CTFs). However, the specific cleavage site(s) and the biochemical properties as well as the functional consequences of pathological TDP-43 CTFs remained unknown. Here we have identified the specific cleavage site, Arg(208), of a pathological TDP-43 CTF purified from FTLD-U brains and show that the expression of this and other TDP-43 CTFs in cultured cells recapitulates key features of TDP-43 proteinopathy. These include the formation of cytoplasmic aggregates that are ubiquitinated and abnormally phosphorylated at sites found in FTLD-U and ALS brain and spinal cord samples. Furthermore, we observed splicing abnormalities in a cell culture system expressing TDP-43 CTFs, and this is significant because the regulation of exon splicing is a known function of TDP-43. Thus, our results show that TDP-43 CTF expression recapitulates key biochemical features of pathological TDP-43 and support the hypothesis that the generation of TDP-43 CTFs is an important step in the pathogenesis of FTLD-U and ALS.

Improved Identification of Endogenous Peptides from Murine Nervous Tissue by Multiplexed Peptide Extraction Methods and Multiplexed Mass Spectrometric Analysis

In recent years, mass spectrometry (MS) based techniques have made their entrance in the analysis of endogenous peptides extracted from nervous tissue. In this study, we introduce a novel peptide extraction procedure using 8 M urea, next to the more established extraction method that uses acetic acid. The extracted peptide mixtures are analyzed by both high-resolution nanoLC MS/MS using collision induced dissociation (CID) on an LTQ-Orbitrap and nanoLC electron transfer induced dissociation (ETD) on a linear ion trap. The combined use of the two extraction methods significantly increased the yield of identified endogenous neuropeptides. The multiplexed use of high mass accuracy mass spectrometry and the ETD fragmentation technique further increased the yield and confidence of peptide identifications. Furthermore, reduction of disulfide bridges during sample preparation was essential in the identification of several endogenous peptides containing cysteine disulfide bonds. Through this study, we identified in total 142 peptides in extracts of the mouse pituitary tissue, whereby 43 uniquely in the urea extract and 11 uniquely in the acetic acid extract. A large number of detected endogenous peptides were reported previously, but we confidently identified 22 unreported peptides that possess characteristics of endogenous peptides and are thus interesting targets to be explored further.

Effect of Biotic and Abiotic Factors on In Vitro Proliferation, Encystment, and Excystment of Pfiesteria piscicida

Pfiesteria spp. are mixotrophic armored dinoflagellates populating the Atlantic coastal waters of the United States. They have been a focus of intense research due to their reported association with several fish mortality events. We have now used a clonal culture of Pfiesteria piscicida and several new environmental isolates to describe growth characteristics, feeding, and factors contributing to the encystment and germination of the organism in both laboratory and environmental samples. We also discuss applied methods of detection of the different morphological forms of Pfiesteria in environmental samples. In summary, Pfiesteria, when grown with its algal prey, Rhodomonas sp., presents a typical growth curve with lag, exponential, and stationary phases, followed by encystment. The doubling time in exponential phase is about 12 h. The profiles of proliferation under a standard light cycle and in the dark were similar, although the peak cell densities were markedly lower when cells were grown in the dark. The addition of urea, chicken manure, and soil extracts did not enhance Pfiesteria proliferation, but crude unfiltered spent aquarium water did. Under conditions of food deprivation or cold (4 degrees C), Pfiesteria readily formed harvestable cysts that were further analyzed by PCR and scanning electron microscopy. The germination of Pfiesteria cysts in environmental sediment was enhanced by the presence of live fish: dinospores could be detected 13 to 15 days earlier and reached 5- to 10-times-higher peak cell densities with live fish than with artificial seawater or f/2 medium alone. The addition of ammonia, urea, nitrate, phosphate, or surprisingly, spent fish aquarium water had no effect.

Modulation of Androgen Receptor Activation Function 2 by Testosterone and Dihydrotestosterone

The androgen receptor (AR) is transcriptionally activated by high affinity binding of testosterone (T) or its 5alpha-reduced metabolite, dihydrotestosterone (DHT), a more potent androgen required for male reproductive tract development. The molecular basis for the weaker activity of T was investigated by determining T-bound ligand binding domain crystal structures of wild-type AR and a prostate cancer somatic mutant complexed with the AR FXXLF or coactivator LXXLL peptide. Nearly identical interactions of T and DHT in the AR ligand binding pocket correlate with similar rates of dissociation from an AR fragment containing the ligand binding domain. However, T induces weaker AR FXXLF and coactivator LXXLL motif interactions at activation function 2 (AF2). Less effective FXXLF motif binding to AF2 accounts for faster T dissociation from full-length AR. T can nevertheless acquire DHT-like activity through an AR helix-10 H874Y prostate cancer mutation. The Tyr-874 mutant side chain mediates a new hydrogen bonding scheme from exterior helix-10 to backbone protein core helix-4 residue Tyr-739 to rescue T-induced AR activity by improving AF2 binding of FXXLF and LXXLL motifs. Greater AR AF2 activity by improved core helix interactions is supported by the effects of melanoma antigen gene protein-11, an AR coregulator that binds the AR FXXLF motif and targets AF2 for activation. We conclude that T is a weaker androgen than DHT because of less favorable T-dependent AR FXXLF and coactivator LXXLL motif interactions at AF2.

Assessment of the “Skin Reservoir” of Urea by Confocal Raman Microspectroscopy and Reverse Iontophoresis in vivo

To demonstrate the "skin reservoir" of urea by confocal Raman microspectroscopy in vivo and to evaluate its impact on the non-invasive monitoring of the analyte by reverse iontophoresis. Urea was extracted iontophoretically over a 2-h period across the skin of adult volunteers and patients with chronic kidney disease. Confocal Raman microspectroscopic profiles of skin were recorded before and after 30 min of current application. Urea extraction was higher at the beginning of current passage, but then decreased to achieve stable values after 2 h of iontophoresis. After 30 min of iontophoresis, the Raman spectra highlighted a clear depletion of urea at the surface of the skin. Lactate distribution was also modified both at the surface and deeper into the skin. A source of urea in the skin, unrelated to the concentration circulating in the blood, was strongly suggested by extracted urea flux observed over time and by the Raman spectroscopy. This "urea reservoir" must be removed before systemic urea levels can be non-invasively monitored by reverse iontophoresis.

In vitro association between the helper component–proteinase of zucchini yellow mosaic virus and cuticle proteins of Myzus persicae

Potyviruses, as typical non-persistently transmitted viruses, are carried within the stylets of aphids. Cuticle proteins (CuPs), which are a major component of the insect cuticle, were examined for in vitro binding to the potyviral helper component-proteinase (HC-Pro). Proteins in 8 M urea extracts from Myzus persicae were separated by SDS-PAGE, electroblotted onto membranes and identified as CuPs by using specific antibodies to M. persicae CuP. Blotted M. persicae protein extracts were overlaid with two HC-Pros, differing by the presence of K or E in the KLSC domain. The HC-Pro with KLSC, known to assist transmission, was found to bind M. persicae proteins, whereas the HC-Pro with ELSC, being deficient in assisting transmission, did not. To identify CuPs that react with HC-Pro, protein extracts were separated by two-dimensional gel electrophoresis. Nine proteins reacting with HC-Pro were sequenced by mass spectrometry. Sequences of peptides in four proteins, of molecular masses between 22 and 31 kDa, were identified as CuPs according to comparison with sequences in GenBank. The putative CuPs from M. persicae that bind HC-Pro are potentially of interest in locating receptors for virions bound to HC-Pro in aphids' stylets.

Segments Crucial for Membrane Translocation and Pore-forming Activity of Bordetella Adenylate Cyclase Toxin

Bordetella adenylate cyclase toxin-hemolysin (CyaA, AC-Hly, or ACT) permeabilizes cell membranes by forming small cation-selective (hemolytic) pores and subverts cellular signaling by delivering into host cells an adenylate cyclase (AC) enzyme that converts ATP to cAMP. Both AC delivery and pore formation were previously shown to involve a predicted amphipathic alpha-helix(502-522) containing a pair of negatively charged Glu(509) and Glu(516) residues. Another predicted transmembrane alpha-helix(565-591) comprises a Glu(570) and Glu(581) pair. We examined the roles of these glutamates in the activity of CyaA. Substitutions of Glu(516) increased specific hemolytic activity of CyaA by two different molecular mechanisms. Replacement of Glu(516) by positively charged lysine residue (E516K) increased the propensity of CyaA to form pores, whereas proline (E516P) or glutamine (E516Q) substitutions extended the lifetime of open single pore units. All three substitutions also caused a drop of pore selectivity for cations. Substitutions of Glu(570) and Glu(581) by helix-breaking proline or positively charged lysine residue reduced (E570K, E581P) or ablated (E570P, E581K) AC membrane translocation. Moreover, E570P, E570K, and E581P substitutions down-modulated also the specific hemolytic activity of CyaA. In contrast, the E581K substitution enhanced the hemolytic activity of CyaA 4 times, increasing both the frequency of formation and lifetime of toxin pores. Negative charge at position 570, but not at position 581, was found to be essential for cation selectivity of the pore, suggesting a role of Glu(570) in ion filtering inside or close to pore mouth. The pairs of glutamate residues in the predicted transmembrane segments of CyaA thus appear to play a key functional role in membrane translocation and pore-forming activities of CyaA.

Phosphorylation of Ser-129 Is the Dominant Pathological Modification of α-Synuclein in Familial and Sporadic Lewy Body Disease

A comprehensive, unbiased inventory of synuclein forms present in Lewy bodies from patients with dementia with Lewy bodies was carried out using two-dimensional immunoblot analysis, novel sandwich enzyme-linked immunosorbent assays with modification-specific synuclein antibodies, and mass spectroscopy. The predominant modification of alpha-synuclein in Lewy bodies is a single phosphorylation at Ser-129. In addition, there is a set of characteristic modifications that are present to a lesser extent, including ubiquitination at Lys residues 12, 21, and 23 and specific truncations at Asp-115, Asp-119, Asn-122, Tyr-133, and Asp-135. No other modifications are detectable by tandem mass spectrometry mapping, except for a ubiquitous N-terminal acetylation. Small amounts of Ser-129 phosphorylated and Asp-119-truncated alpha-synuclein are present in the soluble fraction of both normal and disease brains, suggesting that these Lewy body-associated forms are produced during normal metabolism of alpha-synuclein. In contrast, ubiquitination is only detected in Lewy bodies and is primarily present on phosphorylated synuclein; it therefore likely occurs after phosphorylated synuclein has deposited into Lewy bodies. This invariant pattern of specific phosphorylation, truncation, and ubiquitination is also present in the detergent-insoluble fraction of brain from patients with familial Parkinson's disease (synuclein A53T mutation) as well as multiple system atrophy, suggesting a common pathogenic pathway for both genetic and sporadic Lewy body diseases. These observations are most consistent with a model in which preferential accumulation of normally produced Ser-129 phosphorylated alpha-synuclein is the key event responsible for the formation of Lewy bodies in various Lewy body diseases.

Urea extraction buffer

Urea extraction buffer

Composition of Perineuronal Net Extracellular Matrix in Rat Brain: A DIFFERENT DISACCHARIDE COMPOSITION FOR THE NET-ASSOCIATED PROTEOGLYCANS

We developed a method to extract differentially chondroitin sulfate proteoglycans (CSPGs) that are diffusely present in the central nervous system (CNS) matrix and CSPGs that are present in the condensed matrix of perineuronal nets (PNNs). Adult rat brain was sequentially extracted with Tris-buffered saline (TBS), TBS-containing detergent, 1 m NaCl, and 6 m urea. Extracting tissue sections with these buffers showed that the diffuse and membrane-bound CSPGs were extracted in the first three buffers, but PNN-associated CSPGs remained and were only removed by 6 m urea. Most of the CSPGs were extracted to some degree with all the buffers, with neurocan, brevican, aggrecan, and versican particularly associated with the stable urea-extractable PNNs. The CSPGs in stable complexes only extractable in urea buffer are found from postnatal day 7-14 coinciding with PNN formation. Disaccharide composition analysis indicated a different glycosaminoglycan (GAG) composition for PGs strongly associated with extracellular matrix (ECM). For CS/dermatan sulfate (DS)-GAG the content of nonsulfated, 6-O-sulfated, 2,6-O-disulfated, and 4,6-O-disulfated disaccharides were higher and for heparan sulfate (HS)-GAG, the content of 6-O-sulfated, 2-N-, 6-O-disulfated, 2-O-, 2-N-disulfated, and 2-O-, 2-N-, 6-O-trisulfated disaccharides were higher in urea extract compared with other buffer extracts. Digestions with chondroitinase ABC and hyaluronidase indicated that aggrecan, versican, neurocan, brevican, and phosphacan are retained in PNNs through binding to hyaluronan (HA). A comparison of the brain and spinal cord ECM with respect to CSPGs indicated that the PNNs in both parts of the CNS have the same composition.

Comparison of three methods for enamel protein extraction in different developmental phases of rat lower incisors

Protein extraction methods [urea, trichloroacetic acid (TCA), and acetic acid] were compared for protein recovery from rat incisor developing enamel in the S phase (intermediate/late secretion), M1 phase (early maturation), M2 phase (intermediate maturation), and M3 phase (final maturation). We compared the protein recoveries with the percentage of enamel matrix dry weight burnt off by incineration. Our results indicate that TCA and urea were equally efficient for the extraction of S-stage proteins (85% and 90% recovery, respectively), while urea was the best for M1-stage proteins (92% recovery), and TCA the best for M2-stage (99% recovery) and M3-stage (60% recovery) proteins. The other methods yielded less than 30% recovery in comparison to incineration for M2 and M3 stages. The fact that urea extraction works well in the S and M1 stages and not thereafter is probably related to the changes in the proteins during enamel development and the amount of mineral that needs to be dissolved. TCA is the single method that effectively recovered proteins from all developmental stages of the rat incisor enamel.

High resolution two-dimensional electrophoresis as a tool to differentiate wild from farmed cod ( Gadus morhua) and to assess the protein composition of klipfish

Tris and CHAPS–urea extracts from wild and farmed cod muscle and from rehydrated cod klipfish fillets were analyzed by one (1DE) and two-dimensional electrophoresis (2DE). 2DE maps of tris extracts from farmed cod differed from the wild in a series of spots of Mw 35 and 45 kDa. The CHAPS–urea extracts from farmed cod had a several spots of Mw between 100 and 45 kDa, which were hardly detectable in wild cod and very prominent in klipfish. Klipfish was clearly different from the other samples: the myosin heavy chain was hardly detectable in these samples, and the tris extracts contained fewer, and the CHAPS–urea more spots than the corresponding extracts from the raw muscles. Further identification of these potentially diagnostic spots will make it easier the differentiation of farmed from wild cod and the evaluation of klipfish processing on the protein content of the product.

Multifunctional Glyceraldehyde-3-phosphate Dehydrogenase of Streptococcus pyogenes Is Essential for Evasion from Neutrophils

Streptococcus pyogenes is an important pathogen that causes pharyngitis, sepsis, and rheumatic fever. Cell-associated streptococcal C5a peptidase (ScpA) protects S. pyogenes from phagocytosis and has been suggested to interrupt host defenses by enzymatically cleaving complement C5a, a major factor in the accumulation of neutrophils at sites of infection. How S. pyogenes recognizes and binds to C5a, however, is unclear. We detected a C5a-binding protein in 8 M urea extracts of S. pyogenes by ligand blotting using biotinylated C5a. Searching of genome databases showed that the C5a-binding protein is identical to the streptococcal plasmin receptor (Plr), also known as streptococcal surface dehydrogenase (SDH) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). In the present study we identified a novel function of this multifunctional protein. Western blotting and immunofluorescence microscopy with anti-Plr/SDH/GAPDH showed that Plr/SDH/GAPDH is located on the bacterial surface and released into the culture supernatant. Next, we examined whether the streptococcal Plr/SDH/GAPDH inhibits the biological effects of C5a on human neutrophils. We found that soluble Plr/SDH/GAPDH inhibits C5a-activated chemotaxis and H2O2 production. Furthermore, our results suggested that soluble Plr/SDH/GAPDH captures C5a, inhibiting its chemotactic function. Also, cell-associated Plr/SDH/GAPDH and ScpA were both necessary for the cleavage of C5a on the bacterial surface. Together, these results indicate that the multifunctional protein Plr/SDH/GAPDH has additional functions that help S. pyogenes escape detection by the host immune system.

Analysis of Membrane Proteins from Human Chronic Myelogenous Leukemia Cells: Comparison of Extraction Methods for Multidimensional LC−MS/MS

An important strategy for "shotgun proteomics" profiling involves solution proteolysis of proteins, followed by peptide separation using multidimensional liquid chromatography and automated sequencing by mass spectrometry (LC-MS/MS). Several protocols for extracting and handling membrane proteins for shotgun proteomics experiments have been reported, but few direct comparisons of different protocols have been reported. We compare four methods for preparing membrane proteins from human cells, using acid labile surfactants (ALS), urea, and mixed organic-aqueous solvents. These methods were compared with respect to their efficiency of protein solubilization and proteolysis, peptide and protein recovery, membrane protein enrichment, and peptide coverage of transmembrane proteins. Overall, approximately 50-60% of proteins recovered were membrane-associated, identified from Gene Ontology annotations and transmembrane prediction software. Samples extracted with ALS, extracted with urea followed by dilution, or extracted with urea followed by desalting yielded comparable peptide recoveries and sequence coverage of transmembrane proteins. In contrast, suboptimal proteolysis was observed with organic solvent. Urea extraction followed by desalting may be a particularly useful approach, as it is less costly than ALS and yields satisfactory protein denaturation and proteolysis under conditions that minimize reactivity with urea-derived cyanate. Spectral counting was used to compare datasets of proteins from membrane samples with those of soluble proteins from K562 cells, and to estimate fold differences in protein abundances. Proteins most highly abundant in the membrane samples showed enrichment of integral membrane protein identifications, consistent with their isolation by differential centrifugation.

Matricaria urea extract exhibits antinociceptive activity in male rat

Matricaria urea extract exhibits antinociceptive activity in male rat

Drosophila Lysyl Oxidases Dmloxl-1 and Dmloxl-2 Are Differentially Expressed and the Active DmLOXL-1 Influences Gene Expression and Development

Mammalian lysyl oxidase (LOX) is essential for the catalysis of lysyl-derived cross-links in fibrillar collagens and elastin in the extracellular matrix and has also been implicated in cell motility, differentiation, and tumor cell invasion. The active LOX has been shown to translocate to the nuclei of smooth muscle cells and regulate chromatin structure and transcription. It is difficult to interpret the role of the LOX protein as it is co-expressed with other members of the LOX amine oxidase family in most mammalian cells. To investigate the function of the LOX proteins, we have characterized the Drosophila lysyl oxidases Dmloxl-1 and Dmloxl-2. We present the gene, domain structure, and expression pattern of Dmloxl-1 and Dmloxl-2 during development. In early development, only Dmloxl-1 was expressed, which allowed functional studies. We have expressed Dmloxl-1 in S2 cells and determined that it is a catalytically active enzyme, inhibited by beta-amino-proprionitrile (BAPN), a specific LOX inhibitor. We localized DmLOXL-1 in the nuclei in embryos and in adult salivary gland cells in the nuclei, cytoplasm, and cell surface, using immunostaining and a DmLOXL-1 antibody. To address the biological function of Dmloxl-1, we raised larvae under BAPN inhibitory conditions and over-expressed Dmloxl-1 in transgenic Drosophila. DmLOXL-1 inhibition resulted in developmental delay and a shift in sex ratio; over-expression in the w(m4) variegating strain increased drosopterin production, demonstrating euchromatinization. Our previous data on the transcriptional down-regulation of seven ribosomal genes and the glue gene under inhibitory conditions and the current results collectively support a nuclear role for Dmloxl-1 in euchromatinization and gene regulation.