The cardiac translational landscape reveals that micropeptides are new players involved in cardiomyocyte hypertrophy.

Hypertrophic growth of cardiomyocytes is one of the major compensatory responses in the heart after physiological or pathological stimulation. Protein synthesis enhancement, which is mediated by the translation of messenger RNAs, is one of the main features of cardiomyocyte hypertrophy. Although the transcriptome shift caused by cardiac hypertrophy induced by different stimuli has been extensively investigated, translatome dynamics in this cellular process has been less studied. Here, we generated a nucleotide-resolution translatome as well as transcriptome data from isolated primary cardiomyocytes undergoing hypertrophy. More than 10,000 open reading frames (ORFs) were detected from the deep sequencing of ribosome-protected fragments (Ribo-seq), which orchestrated the shift of the translatome in hypertrophied cardiomyocytes. Our data suggest that rather than increase the translational rate of ribosomes, the increased efficiency of protein synthesis in cardiomyocyte hypertrophy was attributable to an increased quantity of ribosomes. In addition, more than 100 uncharacterized short ORFs (sORFs) were detected in long noncoding RNA genes from Ribo-seq with potential of micropeptide coding. In a random test of 15 candidates, the coding potential of 11 sORFs was experimentally supported. Three micropeptides were identified to regulate cardiomyocyte hypertrophy by modulating the activities of oxidative phosphorylation, the calcium signaling pathway, and the mitogen-activated protein kinase (MAPK) pathway. Our study provides a genome-wide overview of the translational controls behind cardiomyocyte hypertrophy and demonstrates an unrecognized role of micropeptides in cardiomyocyte biology.

Small Molecule Epigenetic Modulators in Pure Chemical Cell Fate Conversion.

Although innovative technologies for somatic cell reprogramming and transdifferentiation provide new strategies for the research of translational medicine, including disease modeling, drug screening, artificial organ development, and cell therapy, recipient safety remains a concern due to the use of exogenous transcription factors during induction. To resolve this problem, new induction approaches containing clinically applicable small molecules have been explored. Small molecule epigenetic modulators such as DNA methylation writer inhibitors, histone methylation writer inhibitors, histone acylation reader inhibitors, and histone acetylation eraser inhibitors could overcome epigenetic barriers during cell fate conversion. In the past few years, significant progress has been made in reprogramming and transdifferentiation of somatic cells with small molecule approaches. In the present review, we systematically discuss recent achievements of pure chemical reprogramming and transdifferentiation.

Direct Reprogramming of Mouse Fibroblasts to Neural Stem Cells by Small Molecules.

Although it is possible to generate neural stem cells (NSC) from somatic cells by reprogramming technologies with transcription factors, clinical utilization of patient-specific NSC for the treatment of human diseases remains elusive. The risk hurdles are associated with viral transduction vectors induced mutagenesis, tumor formation from undifferentiated stem cells, and transcription factors-induced genomic instability. Here we describe a viral vector-free and more efficient method to induce mouse fibroblasts into NSC using small molecules. The small molecule-induced neural stem (SMINS) cells closely resemble NSC in morphology, gene expression patterns, self-renewal, excitability, and multipotency. Furthermore, the SMINS cells are able to differentiate into astrocytes, functional neurons, and oligodendrocytes in vitro and in vivo. Thus, we have established a novel way to efficiently induce neural stem cells (iNSC) from fibroblasts using only small molecules without altering the genome. Such chemical induction removes the risks associated with current techniques such as the use of viral vectors or the induction of oncogenic factors. This technique may, therefore, enable NSC to be utilized in various applications within clinical medicine.

The intracellular domain of sortilin interacts with amyloid precursor protein and regulates its lysosomal and lipid raft trafficking.

The processing of Amyloid precursor protein (APP) is multifaceted, comprising of protein transport, internalization and sequential proteolysis. However, the exact mechanism of APP intracellular trafficking and distribution remains unclear. To determine the interaction between sortilin and APP and the effect of sortilin on APP trafficking and processing, we studied the binding site and its function by mapping experiments, colocalization, coimmunoprecipitation and sucrose gradient fractionation. We identified for the first time that sortilin interacts with APP at both N- and C-terminal regions. The sortilin-FLVHRY (residues 787-792) and APP-NPTYKFFE (residues 759-766) motifs are crucial for the C-terminal interaction. We also found that lack of the FLVHRY motif reduces APP lysosomal targeting and increases APP distribution in lipid rafts in co-transfected HEK293 cells. These results are consistent with our in vivo data where sortilin knockout mice showed a decrease of APP lysosomal distribution and an increase of APP in lipid rafts. We further confirmed that overexpression of sortilin-FLVHRY mutants failed to rescue the lysosomal degradation of APP. Thus, our data suggests that sortilin is implicated in APP lysosomal and lipid raft targeting via its carboxyl-terminal F/YXXXXF/Y motif. Our study provides new molecular insights into APP trafficking and processing.

An optimal method of DNA silver staining in polyacrylamide gels.

DNA silver staining has widely been used to detect DNA fragments in polyacrylamide gels with high sensitivity. We developed an optimal method for DNA silver staining on polyacrylamide gels. The novel procedure can be completed within 10 min instead of over 20 min with the conventional methods. The sensitivity is significantly improved by the silver-ion sensitizer (Eriochrome black T (EBT)) and the minimum of 0.11 and 1.75 ng of DNA amount can be detected in denaturing and nondenaturing polyacrylamide gel, respectively. Compared with the conventional silver staining methods, the improved optimal method can save time and display high sensitivity, color uniformity, and long storage time of the staining gels.