Nuclear envelope localization of PIG-B is essential for GPI-anchor synthesis in Drosophila.

Membrane lipid biosynthesis is a complex process that takes place in various intracellular compartments. Glycosylphosphatidylinositol (GPI), a lipid involved in membrane anchoring of some proteins, is synthesized by the PIG enzymes. Most PIGs are localized to the endoplasmic reticulum (ER), but Drosophila PIG-B (DmPIG-B) is localized to the nuclear envelope (NE). To determine whether the NE localization of DmPIG-B is functionally important, we defined the determinants of localization and generated an ER-localized form, denoted DmPIG-B[ER]. The enzymatic activity of DmPIG-B[ER] was comparable to that of NE-localized DmPIG-B[NE]. Expression of DmPIG-B[ER] inefficiently rescued the lethality of the PIG-B mutant, whereas DmPIG-B[NE] rescued this lethality fully. DmPIG-B[ER] was preferentially degraded by lysosomes, suggesting that the NE localization is essential for function and stability of the protein. In addition, we found that the region of the ER proximal to the NE is the site of translation of GPI-anchored proteins and addition of GPI. Thus, the NE and proximal ER may provide a platform for efficient GPI anchoring.

A neural cell automated analysis system based on pathological specimens in a gerbil brain ischemia model.

PURPOSE: Amid rising health awareness, natural products which has milder effects than medical drugs are becoming popular. However, only few systems can quantitatively assess their impact on living organisms. Therefore, we developed a deep-learning system to automate the counting of cells in a gerbil model, aiming to assess a natural product's effectiveness against ischemia. METHODS: The image acquired from paraffin blocks containing gerbil brains was analyzed by a deep-learning model (fine-tuned Detectron2). RESULTS: The counting system achieved a 79%-positive predictive value and 85%-sensitivity when visual judgment by an expert was used as ground truth. CONCLUSIONS: Our system evaluated hydrogen water's potential against ischemia and found it potentially useful, which is consistent with expert assessment. Due to natural product's milder effects, large data sets are needed for evaluation, making manual measurement labor-intensive. Hence, our system offers a promising new approach for evaluating natural products.

Evaluation and application of RNA-Seq by MinION.

The current RNA-Seq method analyses fragments of mRNAs, from which it is occasionally difficult to reconstruct the entire transcript structure. Here, we performed and evaluated the recent procedure for full-length cDNA sequencing using the Nanopore sequencer MinION. We applied MinION RNA-Seq for various applications, which would not always be easy using the usual RNA-Seq by Illumina. First, we examined and found that even though the sequencing accuracy was still limited to 92.3%, practically useful RNA-Seq analysis is possible. Particularly, taking advantage of the long-read nature of MinION, we demonstrate the identification of splicing patterns and their combinations as a form of full-length cDNAs without losing precise information concerning their expression levels. Transcripts of fusion genes in cancer cells can also be identified and characterized. Furthermore, the full-length cDNA information can be used for phasing of the SNPs detected by WES on the transcripts, providing essential information to identify allele-specific transcriptional events. We constructed a catalogue of full-length cDNAs in seven major organs for two particular individuals and identified allele-specific transcription and splicing. Finally, we demonstrate that single-cell sequencing is also possible. RNA-Seq on the MinION platform should provide a novel approach that is complementary to the current RNA-Seq.