Erratum: Dendritic spine formation and synapse maturation in transcription factor-induced human iPSC-derived neurons.

[This corrects the article DOI: 10.1016/j.isci.2023.106285.].

Dendritic spine formation and synapse maturation in transcription factor-induced human iPSC-derived neurons.

Synaptic maturation is reportedly limited in human induced pluripotent stem cell (iPSC)-derived neurons. Notably, their ability to reach postnatal-like stages and form dendritic spines has been difficult to demonstrate unless using long-term cultured organoids. Recent transcription factor (TF)-based induction methods allow the accelerated generation of differentiated neurons, which offers an unprecedented opportunity to address further progression into late developmental stages. Herein, we report on a comprehensive time-course study of TF-induced iPSC neurons cultured in vitro through an intrinsic maturation program following neurogenesis. Moreover, we determined the transcriptional and morphological sequences of key developmental events associated with spinogenesis, including the conversion of drebrin to its brain-specific isoform A and the N-methyl-D-aspartate (NMDA) receptor subunit switch. TF-induced iPSC neurons successfully acquired structural and functional synaptic maturity, which will critically expand their utility in modeling higher brain functions and disorders.

Overexpression profiling reveals cellular requirements in the context of genetic backgrounds and environments.

Overexpression can help life adapt to stressful environments, making an examination of overexpressed genes valuable for understanding stress tolerance mechanisms. However, a systematic study of genes whose overexpression is functionally adaptive (GOFAs) under stress has yet to be conducted. We developed a new overexpression profiling method and systematically identified GOFAs in Saccharomyces cerevisiae under stress (heat, salt, and oxidative). Our results show that adaptive overexpression compensates for deficiencies and increases fitness under stress, like calcium under salt stress. We also investigated the impact of different genetic backgrounds on GOFAs, which varied among three S. cerevisiae strains reflecting differing calcium and potassium requirements for salt stress tolerance. Our study of a knockout collection also suggested that calcium prevents mitochondrial outbursts under salt stress. Mitochondria-enhancing GOFAs were only adaptive when adequate calcium was available and non-adaptive when calcium was deficient, supporting this idea. Our findings indicate that adaptive overexpression meets the cell's needs for maximizing the organism's adaptive capacity in the given environment and genetic context.

N-terminal deletion of Swi3 created by the deletion of a dubious ORF YJL175W mitigates protein burden effect in S. cerevisiae.

Extreme overproduction of gratuitous proteins can overload cellular protein production resources, leading to growth defects, a phenomenon known as the protein burden/cost effect. Genetic screening in the budding yeast Saccharomyces cerevisiae has isolated several dubious ORFs whose deletions mitigated the protein burden effect, but individual characterization thereof has yet to be delineated. We found that deletion of the YJL175W ORF yielded an N-terminal deletion of Swi3, a subunit of the SWI/SNF chromatin remodeling complex, and partial loss of function of Swi3. The deletion mutant showed a reduction in transcription of genes encoding highly expressed, secreted proteins and an overall reduction in translation. Mutations in the chromatin remodeling complex could thus mitigate the protein burden effect, likely by reallocating residual cellular resources used to overproduce proteins. This cellular state might also be related to cancer cells, as they frequently harbor mutations in the SWI/SNF complex.

Why and how to study genetic changes with context-dependent effects.

The phenotypic impacts of a genetic change can depend on genetic background (e.g. epistasis), as well as other contexts including environment, developmental stage, cell type, disease state, and higher-order combinations thereof. Recent advances in high-throughput phenotyping are uncovering examples of context dependence faster than genotype-phenotype maps and other core concepts are changing to reflect the dynamic nature of biological systems. Here, we review several approaches to study context dependence and their findings. In our opinion, these findings encourage more studies that examine the spectrum of effects a genetic change may have, as opposed to studies that exclusively measure the impact of a genetic change in a particular context. Studies that elucidate the mechanisms that cause the effects of genetic change to vary with context are of special interest. Previous studies of the mechanisms underlying context dependence have improved predictions of phenotype from genotype and have provided insight about how biological systems function and evolve.

A unique Zn(II)2-Cys6-type protein, KpeA, is involved in secondary metabolism and conidiation in Aspergillus oryzae.

Aspergillus oryzae is an important microorganism in the bio- and food industries; therefore, understanding the mechanism underlying its secondary metabolism regulation is important for ensuring its safe use. Here, we screened a novel Zn(II)2-Cys6-type protein-encoding gene, AO090003001186, designated as kpeA (kojic acid production enhancement A), from an A. oryzae disruption mutant library of transcriptional regulators. kpeA is highly conserved among filamentous fungi and encodes a protein with Zn(II)2-Cys6 motif located in the middle of the sequence. Phylogenetic analysis revealed that KpeA was classified into a distal group compared to other fungal Zn(II)2-Cys6-type transcriptional regulators. A Cys to Ala substitution mutant of KpeA showed identical phenotype to the kpeA disruption strain, confirming that KpeA is novel type Zn(II)2-Cys6 binding protein. Colonies of the kpeA disruption strain (ΔkpeA) had longer aerial hyphae and showed decreased conidia production. Microscopic analysis suggested that the reduced vesicle size and conidial head formation in ΔkpeA strain account for the decreased conidia production. Transcriptional levels of brlA and downstream abaA and wetA were decreased in ΔkpeA strain. Moreover, ΔkpeA strain produced 6-fold more kojic acid than the control strains, and the expression of kojR and kojA was increased in ΔkpeA strain. Therefore, KpeA is a novel Zn(II)2-Cys6-type protein likely involved in conidiation and kojic acid production at the transcriptional level.

Estimating the protein burden limit of yeast cells by measuring the expression limits of glycolytic proteins.

The ultimate overexpression of a protein could cause growth defects, which are known as the protein burden. However, the expression limit at which the protein-burden effect is triggered is still unclear. To estimate this limit, we systematically measured the overexpression limits of glycolytic proteins in Saccharomyces cerevisiae. The limits of some glycolytic proteins were up to 15% of the total cellular protein. These limits were independent of the proteins' catalytic activities, a finding that was supported by an in silico analysis. Some proteins had low expression limits that were explained by their localization and metabolic perturbations. The codon usage should be highly optimized to trigger the protein-burden effect, even under strong transcriptional induction. The S-S-bond-connected aggregation mediated by the cysteine residues of a protein might affect its expression limit. Theoretically, only non-harmful proteins could be expressed up to the protein-burden limit. Therefore, we established a framework to distinguish proteins that are harmful and non-harmful upon overexpression.