Growth of Chaetomium cellulolyticum on Alkali-Pretreated Hardwood Sawdust Solids and Pretreatment Liquor

The treatment of a hardwood sawdust with 1% NaOH solution at 121°C dissolved 19.7% of the dry matter, mainly hemicellulose and lignin. Fermentation of the treated solids by Chaetomium cellulolyticum for 48 h gave a product containing 12.5% crude protein (total N × 6.25) on a dry weight basis. The in vitro rumen digestibility of the 48-h fermentation product was 30%, compared to 24% for the alkali-treated but unfermented sawdust. Growth was independent of sawdust particle size in the range 40 to 100 mesh. Fermentation of the pretreatment liquor gave a product containing up to 50% crude protein (dry weight basis) with an in vitro rumen digestibility of 65 to 76%. Approximately 6.7 g of crude protein was obtained from the treated solids and 2.2 g from the pretreatment liquor per 100 g of sawdust treated. The product from the pretreatment liquor fermentation has potential as a high-protein animal feed supplement but could not be produced economically without an outlet for the relatively indigestible product from the solids fermentation. Growth on the pretreatment liquor was strongly pH dependent; there was a considerable increase in the lag phase when the pH was lowered from 7.5 to 5.2. This effect appears to be due to an inhibitor whose toxicity is reduced at high pH.     

The relationship between anogenital distance, fatherhood, and fertility in adult men

Background

Anogenital distance (AGD), a sexually dimorphic measure of genital development, is a marker for endocrine disruption in animal studies and may be shorter in infant males with genital anomalies. Given the correlation between anogenital distance and genital development, we sought to determine if anogenital distance varied in fertile compared to infertile adult men.

Methods

A cross sectional study of consecutive men being evaluated for infertility and men with proven fertility was recruited from an andrology clinic. Anogenital distance (the distance from the posterior aspect of the scrotum to the anal verge) and penile length (PL) were measured using digital calipers. ANOVA and linear regression were used to determine correlations between AGD, fatherhood status, and semen analysis parameters (sperm density, motility, and total motile sperm count).

Findings

A total of 117 infertile men (mean age: 35.3±17.4) and 56 fertile men (mean age: 44.8±9.7) were recruited. The infertile men possessed significantly shorter mean AGD and PL compared to the fertile controls (AGD: 31.8 vs 44.6 mm, PL: 107.1 vs 119.5 mm, p<0.01). The difference in AGD persisted even after accounting for ethnic and anthropomorphic differences. In addition to fatherhood, on both unadjusted and adjusted linear regression, AGD was significantly correlated with sperm density and total motile sperm count. After adjusting for demographic and reproductive variables, for each 1 cm increase in a man''s AGD, the sperm density increases by 4.3 million sperm per mL (95% CI 0.53, 8.09, p = 0.03) and the total motile sperm count increases by 6.0 million sperm (95% CI 1.34, 10.58, p = 0.01). On adjusted analyses, no correlation was seen between penile length and semen parameters.

Conclusion

A longer anogenital distance is associated with fatherhood and may predict normal male reproductive potential. Thus, AGD may provide a novel metric to assess reproductive potential in men.     

Minimal antizyme peptide fully functioning in the binding and inhibition of ornithine decarboxylase and antizyme inhibitor

Antizyme (AZ) is a protein with 228 amino acid residues that regulates ornithine decarboxylase (ODC) by binding to ODC and dissociating its homodimer, thus inhibiting its enzyme activity. Antizyme inhibitor (AZI) is homologous to ODC, but has a higher affinity than ODC for AZ. In this study, we quantified the biomolecular interactions between AZ and ODC as well as AZ and AZI to identify functional AZ peptides that could bind to ODC and AZI and inhibit their function as efficiently as the full-length AZ protein. For these AZ peptides, the inhibitory ability of AZ_95-228 was similar to that of AZ_WT. Furthermore, AZ_95-176 displayed an inhibition (IC(50): 0.20 μM) similar to that of AZ-95-228 (IC(50): 0.16 μM), even though a large segment spanning residues 177-228 was deleted. However, further deletion of AZ_95-176 from either the N-terminus or the C-terminus decreased its ability to inhibit ODC. The AZ_100-176 and AZ_95-169 peptides displayed a noteworthy decrease in ability to inhibit ODC, with IC(50) values of 0.43 and 0.37 μM, respectively. The AZ_95-228, AZ_100-228 and AZ_95-176 peptides had IC(50) values comparable to that of AZ_WT and formed AZ-ODC complexes with K(d,AZ-ODC) values of 1.5, 5.3 and 5.6 μM, respectively. Importantly, our data also indicate that AZI can rescue AZ peptide-inhibited ODC enzyme activity and that it can bind to AZ peptides with a higher affinity than ODC. Together, these data suggest that these truncated AZ proteins retain their AZI-binding ability. Thus, we suggest that AZ_95-176 is the minimal AZ peptide that is fully functioning in the binding of ODC and AZI and inhibition of their function.     

Burkholderia multivorans acts as an antagonist against the growth of Burkholderia pseudomallei in soil

In this study, it was demonstrated, by using agar diffusion tests and a Transwell system, that Burkholderia multivorans NKI379 has an antagonistic effect against the growth of B. pseudomallei. Bacterial representatives were isolated from agricultural crop soil and mixed to construct a partial bacterial community structure that was based on the results of reproducible patterns following PCR-denaturing gradient gel electrophoresis analysis of total soil chromosomes. The antagonistic effect of B. multivorans on B. pseudomallei was observed in this imitate community. In a field study of agricultural crop soil, the presence of B. pseudomallei was inversely related to the presence of the antagonistic strains B. multivorans or B. cenocepacia. B. multivorans NKI379 can survive in a broader range of pH, temperatures and salt concentrations than B. pseudomallei, suggesting that B. multivorans can adapt to extreme environmental changes and therefore predominates over B. pseudomallei in natural environments.     

A novel prenylated C6–C3 compound with estrogen-like activity from the fruits of Illicium arborescens

A novel C₆–C₃ prenylated compound, illicarborene A (1), together with illioliganfunone D (2), 1-allyl-3,5-dimethoxy-4-(3-methylbut-2-enyloxy)benzene (3), (?)-illicinone A (4), (?)-illicinone B (5) and (?)-illicinone A derivative (6) was isolated and characterized from the fruits of Illicium arborescens Hayata. Compound 1 possesses a new class of tricyclic 6/6/5 ring system. The structure of 1 was determined by spectroscopic analysis such as ¹H–¹H COSY, HMQC, HMBC, and NOESY, and confirmed by chemical reaction to yield 7. Compounds 1–5 were found to increase proliferative activity in primary cell culture of osteoblast cells.     

HaplotypeCN: Copy Number Haplotype Inference with Hidden Markov Model and Localized Haplotype Clustering

Copy number variation (CNV) has been reported to be associated with disease and various cancers. Hence, identifying the accurate position and the type of CNV is currently a critical issue. There are many tools targeting on detecting CNV regions, constructing haplotype phases on CNV regions, or estimating the numerical copy numbers. However, none of them can do all of the three tasks at the same time. This paper presents a method based on Hidden Markov Model to detect parent specific copy number change on both chromosomes with signals from SNP arrays. A haplotype tree is constructed with dynamic branch merging to model the transition of the copy number status of the two alleles assessed at each SNP locus. The emission models are constructed for the genotypes formed with the two haplotypes. The proposed method can provide the segmentation points of the CNV regions as well as the haplotype phasing for the allelic status on each chromosome. The estimated copy numbers are provided as fractional numbers, which can accommodate the somatic mutation in cancer specimens that usually consist of heterogeneous cell populations. The algorithm is evaluated on simulated data and the previously published regions of CNV of the 270 HapMap individuals. The results were compared with five popular methods: PennCNV, genoCN, COKGEN, QuantiSNP and cnvHap. The application on oral cancer samples demonstrates how the proposed method can facilitate clinical association studies. The proposed algorithm exhibits comparable sensitivity of the CNV regions to the best algorithm in our genome-wide study and demonstrates the highest detection rate in SNP dense regions. In addition, we provide better haplotype phasing accuracy than similar approaches. The clinical association carried out with our fractional estimate of copy numbers in the cancer samples provides better detection power than that with integer copy number states.     

Influence of sequence and distance between two operators on interaction with the lac repressor

The influence of additional operator or pseudooperator sequences on the lactose repressor-operator interaction has been investigated. Results of kinetic and equilibrium binding measurements suggest an important in vivo role for the Z-gene pseudooperator in repressor-operator binding; the formation of a ternary, looped complex is indicated by the influence of secondary operator sites on binding parameters. Although the binding affinity of the Z-gene pseudooperator [Oz] is only approximately 1/30 that observed for the primary operator [O], the binding affinity to DNA containing both Oz and O is significantly higher than either sequence alone or the sum of the two. This synergistic effect is enhanced further by replacing the pseudooperator sequence [Oz] with the primary operator sequence and results in an even stronger ternary complex in plasmids with duplicate primary sites. The distance between the center of the two primary operators affects the formation of a ternary complex in the linear DNA molecules. Decreased dissociation rate constants were observed with spacing of operator-like sequences between 300 and 500 base pairs (bp). Minimal influence of the second operator on repressor binding is observed when the operators are separated by approximately 4000 or approximately 100 bp. The significant influence of distance on kinetic and equilibrium parameters was demonstrated by measurements on plasmid pRW1511 [Oi-O-PL-Oz] cleaved with restriction enzymes either in the polylinker region to place Oi-O and Oz on opposite ends of the linear plasmid or outside this region to maintain the sites within 500 bp. These results are consistent with the formation of operator-repressor-pseudooperator ternary complex to generate a looped DNA structure.     

Influence of supercoiling and sequence context on operator DNA binding with lac repressor

The dissociation of the repressor-operator complex from a series of negatively supercoiled plasmid DNAs was examined as a function of the sequence context, orientation, and spacing. The plasmids were grouped into four classes, each with common sequence context. The highest dissociation rate constants were observed for the plasmids containing only a single operator (or pseudooperator) sequence, while approximately 10-fold lower rate constants were measured for plasmids with the I gene pseudooperator in conjunction with either the Z gene pseudooperator or the primary operator. Comparison of the behavior of these two classes of plasmids demonstrated the importance of two operator sequences and supported a model of DNA loop formation to stabilize the repressor-operator complex (Whitson, P. A., and Matthews, K. S. (1986) Biochemistry 25, 3845-3852; Whitson, P. A., Olson, J. S., and Matthews, K. S. (1986) Biochemistry 25, 3852-3858; Whitson, P. A., Hsieh, W. T., Wells, R. D., and Matthews, K. S. (1987) J. Biol. Chem. 262, 4943-4946; Kr?mer, H., Niem?ller, M., Amouyal, M., Revet, B., von Wilcken-Bergmann, B., and Müller-Hill, B. (1987) EMBO J. 6, 1481-1491). The third class, with intermediate dissociation rate constants, was comprised of plasmids which contained the primary operator and the higher affinity pseudooperator normally located in the Z gene. Neither the additional presence of the I gene pseudooperator nor the orientation of the primary operator relative to the Z gene pseudooperator significantly affected the dissociation rate constants. The binding characteristics of this group of plasmids demonstrated the essential role of the Z gene pseudooperator in the formation of intramolecular ternary complex and suggested an in vivo function for this pseudooperator. Plasmids containing two primary operator sequences were the class with lowest dissociation rate constants from lac repressor, and minimal effects of salt or spacing on dissociation of this class were observed. These data are consistent with formation of an intramolecular complex with a looped DNA segment stabilized by the combination of increased local concentration of binding sites and torsional stresses on the DNA which favor binding in supercoiled DNA.     

Full-length expanded ataxin-3 enhances mitochondrial-mediated cell death and decreases Bcl-2 expression in human neuroblastoma cells

Machado-Joseph disease (MJD) is an autosomal dominant spinocerebellar degeneration characterized by a wide range of clinical manifestations. An unstable CAG trinucleotide repeat expansion in MJD gene on long arm of chromosome 14 has been identified as the pathologic mutation of MJD and apoptosis was previously shown to be responsible for the neuronal cell death of the disease. In this study, we utilized human neuronal SK-N-SH cells stably transfected with HA-tagged full-length MJD with 78 polyglutamine repeats to examine the effects of polyglutamine expansion on neuronal cell survival in the early stage of disease. Various pro-apoptotic agents were used to assess the tolerance of the mutant cells and to compare the differences between cells with and without mutant ataxin-3. Concentration- and time-dependent experiments showed that the increase in staurosporine-induced cell death was more pronounced and accelerated in cells containing expanded ataxin-3 via MTS assays. Interestingly, under basal conditions, Western blot and immunocytochemical analyses showed a significant decrease of Bcl-2 protein expression and an increase of cytochrome c in cells containing expanded ataxin-3 when compared with those of the parental cells. The same reduction of Bcl-2 was further confirmed in fibroblast cells with mutant ataxin-3. In addition, exogenous expression of Bcl-2 desensitized SK-N-SH-MJD78 cells to poly-Q toxicity. These results indicated that mitochondrial-mediated cell death plays a role in the pathogenesis of MJD. In our cellular model, full-length expanded ataxin-3 that leads to neurodegenerative disorders significantly impaired the expression of Bcl-2 protein, which may be, at least in part, responsible for the weak tolerance to polyglutamine toxicity at the early stage of disease and ultimately resulted in an increase of stress-induced cell death upon apoptotic stress.     

Epstein-Barr virus BGLF4 kinase retards cellular S-phase progression and induces chromosomal abnormality

Epstein-Barr virus (EBV) induces an uncoordinated S-phase-like cellular environment coupled with multiple prophase-like events in cells replicating the virus. The EBV encoded Ser/Thr kinase BGLF4 has been shown to induce premature chromosome condensation through activation of condensin and topoisomerase II and reorganization of the nuclear lamina to facilitate the nuclear egress of nucleocapsids in a pathway mimicking Cdk1. However, the observation that RB is hyperphosphorylated in the presence of BGLF4 raised the possibility that BGLF4 may have a Cdk2-like activity to promote S-phase progression. Here, we investigated the regulatory effects of BGLF4 on cell cycle progression and found that S-phase progression and DNA synthesis were interrupted by BGLF4 in mammalian cells. Expression of BGLF4 did not compensate Cdk1 defects for DNA replication in S. cerevisiae. Using time-lapse microscopy, we found the fate of individual HeLa cells was determined by the expression level of BGLF4. In addition to slight cell growth retardation, BGLF4 elicits abnormal chromosomal structure and micronucleus formation in 293 and NCP-TW01 cells. In Saos-2 cells, BGLF4 induced the hyperphosphorylation of co-transfected RB, while E2F1 was not released from RB-E2F1 complexes. The E2F1 regulated activities of the cyclin D1 and ZBRK1 promoters were suppressed by BGLF4 in a dose dependent manner. Detection with phosphoamino acid specific antibodies revealed that, in addition to Ser780, phosphorylation of the DNA damage-responsive Ser612 on RB was enhanced by BGLF4. Taken together, our study indicates that BGLF4 may directly or indirectly induce a DNA damage signal that eventually interferes with host DNA synthesis and delays S-phase progression.