Factors influencing efficiency of transient gene expression in the red macrophyte Porphyra yezoensis

The marine red alga Porphyra yezoensis has been proposed as a model plant for physiological and genetic studies in seaweeds because of its biological and economical importance. However, the progress of molecular biological studies using gene transfection and genetic transformation systems has been hindered by difficulties in the expression of foreign genes in P. yezoensis cells. To overcome this situation, we developed a transient gene expression system to monitor gene expression in P. yezoensis cells. An artificial υ-glucuronidase (GUS) coding region was synthesized to adapt it to the codon usage of P. yezoensis (PyGUS) and then evaluated for efficiency as a reporter of transient gene expression by particle bombardment. We also demonstrated the importance of using the promoter of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene from P. yezoensis for efficient expression of PyGUS, because the cauliflower mosaic virus (CaMV) 35S promoter, which has been successfully used for monitoring gene expression in nuclei and chloroplasts of higher plants, was less active in P. yezoensis cells. Therefore, the lack of knowledge about differences in the regulatory machinery of gene expression between P. yezoensis and terrestrial plants seems to be why experimental systems for monitoring gene expression were previously not developed in P. yezoensis. Establishment of the transient gene expression system in P. yezoensis could facilitate biotechnological developments in this organism

De novo gene expression and antisense inhibition in cultured cells of BmTRN-1, cloned from the midgut of the silkworm, Bombyx mori, which is homologous with mammalian TIA-1/R

A cDNA encoding a 388 amino acid TIA-1-type RNA-binding protein (BmTRN-1) was isolated from midgut cDNAs of the silkworm, Bombyx mori, via homologous cloning, in order to characterize its function. The deduced amino acid sequence, most likely encoded by a single copy gene, has significant homology with human TIA-1 and TIAR known as apoptotic regulators and recently reported to function as important factors for either splicing or translation. RT-PCR analysis showed that the BmTRN-1 gene was vigorously transcribed in the midgut at the gut purge stage, indicating a possible relation to the tissue-decomposing process in larval-pupal metamorphosis. We also show that inhibition of the expression of BmTRN-1 by a transfected oligonucleotide encoding the antisense sequence caused a remarkable rise in protein expression from artificially constructed cDNAs encoded by plasmid vectors in Bombyx cells, depending on the constructed ORF sequences of the introduced cDNAs. Furthermore, it was shown that the transcripts from the cDNAs introduced into the cells increased under the antisense-inhibition of BmTRN-1 when the protein levels of these cDNAs also rose, demonstrating that BmTRN-1 could act as a regulator especially of the mechanism eliminating transcripts with possible targets for BmTRN-1 recognition in the authentic post-transcription process

Brain p3-Alcβ peptide restores neuronal viability impaired by Alzheimer's amyloid β-peptide

We propose a new therapeutic strategy for Alzheimer's disease (AD). Brain peptide p3-Alc beta 37 is generated from the neuronal protein alcadein beta through cleavage of gamma-secretase, similar to the generation of amyloid beta (A beta) derived from A beta-protein precursor/APP. Neurotoxicity by A beta oligomers (A beta o) is the prime cause prior to the loss of brain function in AD. We found that p3-Alc beta 37 and its shorter peptide p3-Alc beta 9-19 enhanced the mitochondrial activity of neurons and protected neurons against A beta o-induced toxicity. This is due to the suppression of the A beta o-mediated excessive Ca2+ influx into neurons by p3-Alc beta. Successful transfer of p3-Alc beta 9-19 into the brain following peripheral administration improved the mitochondrial viability in the brain of AD mice model, in which the mitochondrial activity is attenuated by increasing the neurotoxic human A beta 42 burden, as revealed through brain PET imaging to monitor mitochondrial function. Because mitochondrial dysfunction is common in the brain of AD patients alongside increased A beta and reduced p3-Alc beta 37 levels, the administration of p3-Alc beta 9-19 may be a promising treatment for restoring, protecting, and promoting brain functions in patients with AD