Metabolites of Trichoderma longibrachiatum EF5 inhibits soil borne pathogen, Macrophomina phaseolina by triggering amino sugar metabolism.

An endophytic fungal antagonist Trichoderma longibrachiatum EF5 exhibited biocontrol activity against a soil-borne fungal pathogen Macrophomina phaseolina. Under dual co-culture, T. longibrachiatum EF5 showed 58% inhibition against M. phaseolina. Crude soluble metabolites (SMs) extracted from EF5 exhibited biocontrol activity (61%), which is more significant than the cell-free extract. Dual culture of both T. longibrachiatum EF5 and M. phaseolina displayed entangled mycelial structures and retarded hyphal growth. The metabolites responsible for antibiosis and pathogenic activity profiled through GC-MS revealed a total of 131 SMs from axenic culture and upon the interaction of T. longibrachiatum EF5 and M. phaseolina. Interestingly, potential plant-growth-promoting and antimicrobial compounds such as 1- pentanol, 1-hexanol, myristonyl pantothenate, bisabolol, d-Alanine, and diethyl trisulphide were unique with T. longibrachiatum EF5. Few compounds that were not observed or produced minimally under axenic culture were increased during their interaction (e.g., 1,6-anhydro-á-d-Glucopyranose and 5-heptyl dihydro-2(3H)-Furanone), suggesting antimicrobial action against the pathogen. This study also unraveled the induction of amino sugar metabolism when T. longibrachiatum EF5 interacts with M. phaseolina, which is responsible for colonization and counterfeiting the pathogen. Hence T. longibrachiatum EF5 could be a potential biocontrol agent employed for defense priming and plant growth promotion.

Yb:YAG master oscillator power amplifier for remote wind sensing.

We have demonstrated key advances towards a solid-state laser amplifier at 1.03 microm for global remote wind sensing. We designed end-pumped zig-zag slab amplifiers to achieve high gain. We overcame parasitic oscillation limitations using claddings on the slab's total internal reflection (TIR) and edge surfaces to confine the pump and signal light by TIR and allow leakage of amplified spontaneous emission rays that do not meet the TIR condition. This enables e3, e5, and e8 single-, double-, and quadruple-pass small-signal amplifier gain, respectively. The stored energy density is 15.6 J/cm3, a record for a laser-diode end-pumped Yb:YAG zig-zag slab amplifier.

Characterization of CaMKII-expressing neurons within a striatal region implicated in avian vocal learning.

In songbirds, an anterior forebrain pathway has been implicated in vocal learning. Within Area X, the striatal-pallidal component of this forebrain pathway, early social tutoring dramatically increases the autophosphorylation of CaMKII (calcium-calmodulin-dependent protein kinase II). Activation of CaMKII often is associated with forms of synaptic plasticity (e.g. LTP) underlying learning and memory, and electrophysiological studies have demonstrated NMDA and dopamine (DA) receptor-dependent LTP among Area X medium spiny neurons [Ding, L., Perkel, D.J., 2002. Dopamine modulates excitability of spiny neurons in the avian Basal Ganglia. J. Neurosci. 22, 5210-5218]. Together, these data suggest that Area X neurons may help to encode a representation of song used for vocal mimicry. To identify which Area X neurons could participate in the CaMKII response to song tutoring, we used immunocytochemistry to assess the colocalization of CaMKII with several other biochemical markers that identify specific neuron classes within Area X. Virtually all (approximately 98%) Area X cells expressing CaMKII also expressed DARPP-32 (dopamine- and adenosine 3'5'-monophosphate-regulated phosphoprotein), a dopamine signaling protein enriched in medium spiny striatal neurons. The implication that medium spiny neurons are primary mediators of the pCaMKII response to tutoring is interesting in view of the established dopaminergic modulation of LTP in this cell type. Additionally, BrdU and DARPP-32 immunocytochemistry were combined to test whether medium spiny neurons are among the neurons generated and incorporated into Area X during song learning. Based upon their expression of DARPP-32, the majority of Area X neurons labeled by BrdU injections given on posthatch days 20-25 are medium spiny neurons.

A novel gamma -secretase assay based on detection of the putative C-terminal fragment-gamma of amyloid beta protein precursor.

Alzheimer's disease is characterized by the deposits of the 4-kDa amyloid beta peptide (A beta). The A beta protein precursor (APP) is cleaved by beta-secretase to generate a C-terminal fragment, CTF beta, which in turn is cleaved by gamma-secretase to generate A beta. Alternative cleavage of the APP by alpha-secretase at A beta 16/17 generates the C-terminal fragment, CTFalpha. In addition to A beta, endoproteolytic cleavage of CTF alpha and CTF beta by gamma-secretase should yield a C-terminal fragment of 57-59 residues (CTF gamma). However, CTF gamma has not yet been reported in either brain or cell lysates, presumably due to its instability in vivo. We detected the in vitro generation of A beta as well as an approximately 6-kDa fragment from guinea pig brain membranes. We have provided biochemical and pharmacological evidence that this 6-kDa fragment is the elusive CTF gamma, and we describe an in vitro assay for gamma-secretase activity. The fragment migrates with a synthetic peptide corresponding to the 57-residue CTF gamma fragment. Three compounds previously identified as gamma-secretase inhibitors, pepstatin-A, MG132, and a substrate-based difluoroketone (t-butoxycarbonyl-Val-Ile-(S)-4-amino-3-oxo-2, 2-difluoropentanoyl-Val-Ile-OMe), reduced the yield of CTF gamma, providing additional evidence that the fragment arises from gamma-secretase cleavage. Consistent with reports that presenilins are the elusive gamma-secretases, subcellular fractionation studies showed that presenilin-1, CTF alpha, and CTF beta are enriched in the CTF gamma-generating fractions. The in vitro gamma-secretase assay described here will be useful for the detailed characterization of the enzyme and to screen for gamma-secretase inhibitors.

The influence of physical structure and charge on neurite extension in a 3D hydrogel scaffold.

Understanding neural cell differentiation and neurite extension in three-dimensional scaffolds is critical for neural tissue engineering. This study explores the structure-function relationship between a 3D hydrogel scaffold and neural cell process extension and examines the role of ambient charge on neurite extension in 3D scaffolds. A range of agarose hydrogel concentrations was used to generate varied gel physical structures and the corresponding neurite extension was examined. Agarose gel concentration and the corresponding pore radius are important physical properties that influence neural cell function. The average pore radii of the gels were determined while the gel was in the hydrated state and in two different dehydrated states. As the gel concentration was increased, the average pore radius decreased exponentially. Similarly, the length of neurites extended by E9 chick DRGs cultured in agarose gels depends on gel concentration. The polycationic polysaccharide chitosan and the polyanionic polysaccharide alginate were used to incorporate charge into the 3D hydrogel scaffold, and neural cell response to charge was studied. Chitosan and alginate were covalently bound to the agarose hydrogel backbone using the bi-functional coupling agent 1,1'-carbonyldiimidazole. DRGs cultured in chitosan-coupled agarose gel exhibited a significant increase in neurite length compared to the unmodified agarose control. Conversely, the alginate-coupled agarose gels significantly inhibited neurite extension. This study demonstrates a strong, correlation between the ability of sensory ganglia to extend neurites in 3D gels and the hydrogel pore radius. In addition, our results demonstrate that charged biopolymers influence neurite extension in a polarity dependent manner.