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David Page and colleagues report the sequence of the chicken W sex chromosome and compare ancestral W-linked genes across bird species. They find that the W chromosome did not acquire genes expressed exclusively in reproductive tissue, but retained genes through selection to maintain appropriate dosage levels of broadly expressed genes. After birds diverged from mammals, different ancestral autosomes evolved into sex chromosomes in each lineage. In birds, females are ZW and males are ZZ, but in mammals females are XX and males are XY. We sequenced the chicken W chromosome, compared its gene content with our reconstruction of the ancestral autosomes, and followed the evolutionary trajectory of ancestral W-linked genes across birds. Avian W chromosomes evolved in parallel with mammalian Y chromosomes, preserving ancestral genes through selection to maintain the dosage of broadly expressed regulators of key cellular processes. We propose that, like the human Y chromosome, the chicken W chromosome is essential for embryonic viability of the heterogametic sex. Unlike other sequenced sex chromosomes, the chicken W chromosome did not acquire and amplify genes specifically expressed in reproductive tissues. We speculate that the pressures that drive the acquisition of reproduction-related genes on sex chromosomes may be specific to the male germ line.

The human X and Y chromosomes evolved from an ordinary pair of autosomes, but millions of years ago genetic decay ravaged the Y chromosome, and only three per cent of its ancestral genes survived. We reconstructed the evolution of the Y chromosome across eight mammals to identify biases in gene content and the selective pressures that preserved the surviving ancestral genes. Our findings indicate that survival was nonrandom, and in two cases, convergent across placental and marsupial mammals. We conclude that the gene content of the Y chromosome became specialized through selection to maintain the ancestral dosage of homologous X–Y gene pairs that function as broadly expressed regulators of transcription, translation and protein stability. We propose that beyond its roles in testis determination and spermatogenesis, the Y chromosome is essential for male viability, and has unappreciated roles in Turner’s syndrome and in phenotypic differences between the sexes in health and disease. A study comparing the Y chromosome across mammalian species reveals that selection to maintain the ancestral dosage of homologous X–Y gene pairs preserved a handful of genes on the Y chromosome while the rest were lost; the survival of broadly expressed dosage-sensitive regulators of gene expression suggest that the human Y chromosome is essential for male viability. Evolution and function of the Y chromosome Mammalian Y chromosomes, known for their roles in sex determination and male fertility, often contain repetitive sequences that make them harder to assemble than the rest of the genome. To counter this problem Henrik Kaessmann and colleagues have developed a new transcript assembly approach based on male-specific RNA/genomic sequencing data to explore Y evolution across 15 species representing all major mammalian lineages. They find evidence for two independent sex chromosome originations in mammals and one in birds. Their analysis of the Y/W gene repertoires suggests that although some genes evolved novel functions in sex determination/spermatogenesis as a result of temporal/spatial expression changes, most Y genes probably persisted, at least initially, as a result of dosage constraints. In a parallel study, Daniel Bellott and colleagues reconstructed the evolution of the Y chromosome, using a comprehensive comparative analysis of the genomic sequence of X–Y gene pairs from seven placental mammals and one marsupial. They conclude that evolution streamlined the gene content of the human Y chromosome through selection to maintain the ancestral dosage of homologous X–Y gene pairs that regulate gene expression throughout the body. They propose that these genes make the Y chromosome essential for male viability and contribute to differences between the sexes in health and disease.

The male-specific region of rhesus macaque and human Y chromosome (MSY) are sequenced and compared to the human MSY, showing that during the last 25 million years MSY gene loss in the rhesus and human lineages was limited to the youngest stratum (stratum 5), whereas gene loss in the older strata ceased more than 25 million years ago. The fate of the Y chromosome The mammal sex-determining X and Y chromosomes evolved from a pair of autosomes ('normal' non-sex chromosomes), but as a result of genetic decay, the male-specific region of the human Y chromosome retains only 3% of the ancestral autosomal genes. Here, the authors have sequenced the corresponding region of the rhesus macaque Y, and reconstructed the trajectory of its evolution through an analysis of the rhesus, human and chimpanzee Y chromosomes. The results show that initial rapid loss of genes is followed by strict conservation through purifying selection. This runs counter to the view that the human Y chromosome is destined inevitably for extinction. The human X and Y chromosomes evolved from an ordinary pair of autosomes during the past 200–300 million years 1 , 2 , 3 . The human MSY (male-specific region of Y chromosome) retains only three percent of the ancestral autosomes’ genes owing to genetic decay 4 , 5 . This evolutionary decay was driven by a series of five ‘stratification’ events. Each event suppressed X–Y crossing over within a chromosome segment or ‘stratum’, incorporated that segment into the MSY and subjected its genes to the erosive forces that attend the absence of crossing over 2 , 6 . The last of these events occurred 30 million years ago, 5 million years before the human and Old World monkey lineages diverged. Although speculation abounds regarding ongoing decay and looming extinction of the human Y chromosome 7 , 8 , 9 , 10 , remarkably little is known about how many MSY genes were lost in the human lineage in the 25 million years that have followed its separation from the Old World monkey lineage. To investigate this question, we sequenced the MSY of the rhesus macaque, an Old World monkey, and compared it to the human MSY. We discovered that during the last 25 million years MSY gene loss in the human lineage was limited to the youngest stratum (stratum 5), which comprises three percent of the human MSY. In the older strata, which collectively comprise the bulk of the human MSY, gene loss evidently ceased more than 25 million years ago. Likewise, the rhesus MSY has not lost any older genes (from strata 1–4) during the past 25 million years, despite its major structural differences to the human MSY. The rhesus MSY is simpler, with few amplified gene families or palindromes that might enable intrachromosomal recombination and repair. We present an empirical reconstruction of human MSY evolution in which each stratum transitioned from rapid, exponential loss of ancestral genes to strict conservation through purifying selection.

Dynamic sex chromosomes Birds and mammals have distinct sex chromosomes. In birds, males have a pair of Z chromosomes and females a Z and a W. In mammals, males are XY and females XX. It has long been assumed that sex-chromosome evolution has involved dramatic modification of the sex-specific (W and Y) chromosomes but only modest changes to the Z and X chromosomes shared by the sexes. Not so, according to a new study reporting the sequence of the chicken Z chromosome and comparing it with the finished sequence of human X. The Z and X chromosomes have changed dramatically from the autosomal (non-sex) chromosomes that gave rise to them. And they seem to have followed convergent evolutionary trajectories, including the acquisition and amplification of testis-expressed gene families, despite having arisen independently from different portions of the ancestral genome. Birds and mammals have distinct sex chromosomes: in birds, males are ZZ and females ZW; in mammals, males are XY and females XX. By sequencing the chicken Z chromosome and comparing it with the human X chromosome, these authors overturn the currently held view that these chromosomes have diverged little from their autosomal progenitors. The Z and X chromosomes seem to have followed convergent evolutionary trajectories, despite evolving with opposite systems of heterogamety. In birds, as in mammals, one pair of chromosomes differs between the sexes. In birds, males are ZZ and females ZW. In mammals, males are XY and females XX. Like the mammalian XY pair, the avian ZW pair is believed to have evolved from autosomes, with most change occurring in the chromosomes found in only one sex—the W and Y chromosomes 1 , 2 , 3 , 4 , 5 . By contrast, the sex chromosomes found in both sexes—the Z and X chromosomes—are assumed to have diverged little from their autosomal progenitors 2 . Here we report findings that challenge this assumption for both the chicken Z chromosome and the human X chromosome. The chicken Z chromosome, which we sequenced essentially to completion, is less gene-dense than chicken autosomes but contains a massive tandem array containing hundreds of duplicated genes expressed in testes. A comprehensive comparison of the chicken Z chromosome with the finished sequence of the human X chromosome demonstrates that each evolved independently from different portions of the ancestral genome. Despite this independence, the chicken Z and human X chromosomes share features that distinguish them from autosomes: the acquisition and amplification of testis-expressed genes, and a low gene density resulting from an expansion of intergenic regions. These features were not present on the autosomes from which the Z and X chromosomes originated but were instead acquired during the evolution of Z and X as sex chromosomes. We conclude that the avian Z and mammalian X chromosomes followed convergent evolutionary trajectories, despite their evolving with opposite (female versus male) systems of heterogamety. More broadly, in birds and mammals, sex chromosome evolution involved not only gene loss in sex-specific chromosomes, but also marked expansion and gene acquisition in sex chromosomes common to males and females.

Understanding neurodegenerative disease pathology requires a close examination of neurons and their processes. However, image-based single-cell analyses of neurons often require laborious and time-consuming manual classification tasks. Here, we present a machine learning (ML) approach leveraging convolutional neural network (CNN) classifiers capable of accurately identifying various classes of neuronal images, including single neurons. We developed the Single Neuron Identification Model 20-Class (SNIM20) which was trained on a dataset of induced pluripotent stem cell (iPSC)-derived motor neurons, containing over 12,000 images from 20 distinct classes. SNIM20 is built in TensorFlow and trained on images of neurons differentiated from iPSC cultures that were stained for nuclei and microtubules. This classifier demonstrated high predictive accuracy (AUC = 0.99) for distinguishing single neurons. Additionally, the 2-stage training framework can be used more broadly for cellular classification tasks. A variation was successfully trained on images of a human osteosarcoma cell line (U2OS) for single-cell classification (AUC = 0.99). While this framework was primarily designed for single-cell microraft-based identification and capture, it also works with cells in standard plate formats. We additionally explore the impact of fluorescent channels and brightfield images, class groupings, and transfer learning on the quality of the classification. This framework can both assist in high throughput neuronal or cellular identification and be used to train a custom classifier for the user’s specific needs.

Spatial omics (SO) produces high-definition mapping of subcellular molecules within tissue samples. Mapping transcripts to anatomical regions requires segmentation, but this remains challenging in tissue cross-sections with tubular structures like axons in peripheral nerve or spinal cord. Neural networks could address misidentification but are hindered by the need for extensive human annotations. We present SiDoLa-NS (Simulate, Don’t Label-Nervous System), an image-driven (top-down) approach to SO analysis in the nervous system. We utilize biophysical properties of tissue architectures to design synthetic images of tissue samples, eliminating reliance on manual annotation and enabling scalable training data generation. With synthetic samples, we trained supervised instance segmentation convolutional neural networks (CNNs) for nucleus segmentation, achieving precision and F1-scores>0.95. We further identify macroscopic tissue structures in mouse brain (mAP 50 =0.869), spinal cord (mAP 50 =0.96), and pig sciatic nerve (mAP 50 =0.995). This framework sets the stage for transferable models across species and tissue architectures—accelerating SO applications in neuroscience and beyond.

Most human genetic variation is classified as variants of uncertain significance. While advances in genome editing have allowed innovation in pooled screening platforms, many screens deal with relatively simple readouts (viability, fluorescence) and cannot identify the complex cellular phenotypes that underlie most human diseases. In this paper, we present a generalizable functional genomics platform that combines high-content imaging, machine learning, and microraft isolation in a method termed “Raft-Seq”. We highlight the efficacy of our platform by showing its ability to distinguish pathogenic point mutations of the mitochondrial regulator Mitofusin 2, even when the cellular phenotype is subtle. We also show that our platform achieves its efficacy using multiple cellular features, which can be configured on-the-fly. Raft-Seq enables a way to perform pooled screening on sets of mutations in biologically relevant cells, with the ability to physically capture any cell with a perturbed phenotype and expand it clonally, directly from the primary screen. Raft-Seq is a generalizable pooled screening platform that combines high-content imaging, machine learning and microraft isolation, and enables efficient screening of genetic perturbations based on their impact on phenotypes.

Mutations in mitochondrial-related genes underlie numerous neurodegenerative diseases, yet the significance of most variants remains uncertain concerning disease phenotypes. Several thousand genes have been shown to regulate mitochondria in eukaryotic cells, but which of these genes are necessary for proper mitochondrial function and dynamics? We investigated the degree of morphological disruptions in mitochondrial gene-silenced cells to understand the genetic contribution to the expected mitochondrial phenotype and to identify potentially pathogenic variants like pathogenic mutations in MFN2 . We analyzed 5835 gRNAs in a high dimensional phenotypic dataset produced by the image-based pooled analysis platform Raft-Seq. Using the MFN2 -mutant cell phenotype, we identified several genes, including TMEM11 , TIMM8A , NDUFAF4 , NDUFAF7 , and NDUFS5 ( NADH ubiquinone oxidoreductase -related genes), as crucial for normal mitochondrial dynamics in human U2OS cells. Additionally, we found several missense and UTR variants within the genes SLC25A19 and ATAD3A as drivers of mitochondrial aggregation. By examining multiple features instead of a single readout, this analysis was powered to detect genes which had morphological ‘signatures’ aligned with MFN2 -mutant phenotypes. Reanalysis with anomaly detection revealed other critical genes, including APOOL , MCEE , NIT , PHB , and SLC16A7 , which perturb mitochondrial network morphology in a manner divergent from MFN2 . These studies show causal links between gene knockouts and gene-specific variants into the assembly or maintenance of mitochondrial dynamics and can hopefully lead to a better understanding of mitochondrial related diseases.

In mammals, chromosomes occupy defined positions in sperm, whereas previous work in chicken showed random chromosome distribution. Monotremes (platypus and echidnas) are the most basal group of living mammals. They have elongated sperm like chicken and a complex sex chromosome system with homology to chicken sex chromosomes. We used platypus and chicken genomic clones to investigate genome organization in sperm. In chicken sperm, about half of the chromosomes investigated are organized non-randomly, whereas in platypus chromosome organization in sperm is almost entirely non-random. The use of genomic clones allowed us to determine chromosome orientation and chromatin compaction in sperm. We found that in both species chromosomes maintain orientation of chromosomes in sperm independent of random or non-random positioning along the sperm nucleus. The distance of loci correlated with the total length of sperm nuclei, suggesting that chromatin extension depends on sperm elongation. In platypus, most sex chromosomes cluster in the posterior region of the sperm nucleus, presumably the result of postmeiotic association of sex chromosomes. Chicken and platypus autosomes sharing homology with the human X chromosome located centrally in both species suggesting that this is the ancestral position. This suggests that in some therian mammals a more anterior position of the X chromosome has evolved independently.

[...]we have developed a system to track individual regions that are under review. The primary assembly unit contains sequences for the non-redundant haploid assembly; this includes the scaffolds that make up the chromosome sequence as well as unplaced and unlocalized scaffolds that are thought to represent novel sequence (not shown in this picture).\\n Additionally, we wish to engage the research and clinical communities to identify regions that require targeted effort and to incorporate information from groups performing detailed work on specific loci.