Biology subjects

Niakan, K. K.

Publications and source records attributed to Niakan, K. K..

9 recordsLinked to original sources

bioRxiv2026

In situ cryo-ET of mammalian embryos reveals cytoplasmic lattices contain ubiquitin-charged E2-E3 ligase assemblies

Cytoplasmic lattices (CPLs) are filamentous assemblies essential for mammalian embryonic development that regulate organelle organization, spindle assembly, and protein homeostasis. However, their molecular functions remain unclear. Here, we develop a strategy combining cryo-focused ion beam milling and cryo-electron tomography to resolve macromolecular complexes directly in mammalian embryos. Using this approach, we determine the in situ structure of CPLs within mouse embryos at [~]4.7 [A] resolution. CPL filaments are built from multiple copies of fourteen proteins arranged into a [~]4.5 MDa repeating unit. These components form a scaffold that organizes a central cavity containing three complexes of the E2 ubiquitin-conjugating enzyme UBE2D, the E3 ligase UHRF1, NLRP14 and {beta}-tubulin. We observe states in which UBE2D is conjugated to ubiquitin, along with structural rearrangements within the cavity consistent with regulated ubiquitin transfer. Together, our findings suggest that CPLs are large ubiquitin ligase assemblies and play a role in post-translational protein modification during early embryonic development.

molecular biology↗

bioRxiv2026

Basement membrane mechanics drives patterned response to developmental signalling

Precise spatiotemporal patterning of cell fate decisions during development, such as those accompanying gastrulation, has traditionally been attributed to morphogen gradients. Emerging evidence also points to a crucial role for mechanics in regulating cell fate decisions. Here, we uncover that basement membrane mechanics in human pluripotent stem cells (hPSCs) regulates the spatiotemporal response to gastrulation-inducing BMP4. Reducing mechanosensation via soft substrates or chemical inhibitors abolishes stereotypical spatial patterning of the BMP4 response in hPSC colonies. This loss arises from disrupted epithelial polarity and increased permeability, which enhance BMP receptor accessibility. Strikingly, apical exposure to soluble laminin similarly disrupts polarity and suppresses hPSC mechanosensing, phenocopying the soft substrates effect and suggesting feedback between polarity signaling and mechanosensing. Finally, softening the basement membrane in mouse embryos triggers ectopic and premature mesoderm differentiation, disrupting gastrulation patterning. Together, this study describes a mechanochemical feedback mechanism that establishes basement membrane mechanics as a key regulator of developmental patterning.

cell biology↗

bioRxiv2025

Dissecting PADI6 function defines oocyte cytoplasmic lattices as regulatory hubs for fundamental cellular processes

Oocyte cytoplasmic lattices are critical for early embryo development but their composition and function are not fully understood. Mutations in PADI6, an essential component of cytoplasmic lattices, lead to early embryonic developmental arrest and female infertility. To investigate PADI6 function in mRNA storage, global protein levels, and lattice composition during early mammalian development we used single cell transcriptomics and proteomics methods to study two mouse models. Padi6 null mutation resulted in inhibition of embryonic genome activation, defective maternal mRNA degradation, and disruption to protein storage on the cytoplasmic lattices. Distinct developmental phenotypes were observed with a hypomorphic Padi6 mutation. By developing a powerful single cell proteomic fractionation method, we define the cytoplasmic lattice enriched proteome in which we find essential components of another major oocyte-specific compartment, the endolysosomal vesicular assembly (ELVA), suggesting previously unknown interconnections between them. Our findings highlight a critical scaffolding function of PADI6 and implicate cytoplasmic lattices as regulatory hubs for key processes in the oocyte and early embryo, including translation, respiration and protein degradation.

developmental biology↗

bioRxiv2024

ETV4 and ETV5 Orchestrate FGF-Mediated Lineage Specification and Epiblast Maturation during Early Mouse Development

Cell fate decisions in early mammalian embryos are tightly regulated processes crucial for proper development. While FGF signaling plays key roles in early embryo patterning, its downstream effectors remain poorly understood. Our study demonstrates that the transcription factors Etv4 and Etv5 are critical mediators of FGF signaling in cell lineage specification and maturation in mouse embryos. We show that loss of Etv5 compromises primitive endoderm formation at pre-implantation stages. Furthermore, Etv4/5 deficiency delays naive pluripotency exit and epiblast maturation, leading to elevated NANOG and reduced OTX2 expression within the blastocyst epiblast. As a consequence of delayed pluripotency progression, Etv4/5 deficient embryos exhibit anterior visceral endoderm migration defects post-implantation, a process essential for coordinated embryonic patterning and gastrulation initiation. Our results demonstrate the successive roles of these FGF signaling effectors in early lineage specification and embryonic body plan establishment, providing new insights into the molecular control of mammalian development. Summary statementFGF signaling effectors ETV4/5 regulate lineage specification and embryo patterning in mice, affecting primitive endoderm formation and pluripotency exit.

developmental biology↗

bioRxiv2024

Marsupial single-cell transcriptomics provides an atlas of developmental heterochrony

Single-cell transcriptomics has revealed conserved and divergent programmes of organogenesis in mammals, but existing studies have focused on eutherians. Marsupials exhibit short gestation and complete development externally, necessitating accelerated differentiation of anterior features required for locomotion and feeding. As such, they represent a unique outgroup with which to understand temporal shifts in development, known as heterochrony. Here, we generate the first single-cell transcriptomic atlas of gastrulation and early organogenesis in a marsupial, the opossum Monodelphis domestica. We find that anterior prioritisation is achieved by earlier initiation and shorter duration of transcriptional programmes relative to eutherians. The result is uncoupling of transcriptional and morphological progression, revealing unforeseen diversity in the order of developmental sequences in mammals. We uncover novel tissues for which heterochrony has not previously been documented. Our findings indicate that accelerated activation of transcriptional programmes facilitates the rapid growth needed for survival of the marsupial neonate.

developmental biology↗

bioRxiv2024

Suppression of ERK signalling promotes pluripotent epiblast in the human blastocyst

Studies in the mouse demonstrate the importance of fibroblast growth factor (FGF) and extra-cellular receptor tyrosine kinase (ERK) in specification of embryo-fated epiblast and yolk-sac-fated hypoblast cells from uncommitted inner cell mass (ICM) cells prior to implantation. Molecular mechanisms regulating specification of early lineages in human development are comparatively unclear. Here we show that exogenous FGF stimulation leads to expanded hypoblast molecular marker expression, at the expense of the epiblast. Conversely, we show that specifically inhibiting ERK activity leads to expansion of epiblast cells functionally capable of giving rise to naive human pluripotent stem cells. Single-cell transcriptomic analysis indicates that these epiblast cells downregulate FGF signalling and upregulate molecular markers associated with naive pluripotency. Our functional study demonstrates for the first time the molecular mechanisms governing ICM specification in human development, whereby segregation of the epiblast and hypoblast lineages occurs during maturation of the mammalian embryo in an ERK signal-dependent manner.

developmental biology↗

bioRxiv2021

Direct reprogramming of human embryonic to trophoblast stem cells

During the first week of development, human embryos form a blastocyst comprised of an inner cell mass and trophectoderm (TE) cells, the latter of which are progenitors of placental trophoblast. Here we investigated the expression of transcripts in the human TE from early to late blastocyst stages. We identified enrichment of transcription factors GATA2, GATA3, TFAP2C and KLF5 and characterised their protein expression dynamics across TE development. By inducible overexpression and mRNA transfection we determined that these factors, together with MYC, are sufficient to establish induced trophoblast stem cells (iTSCs) from primed human embryonic stem cells. These iTSCs self-renew and recapitulate morphological characteristics, gene expression profiles, and directed differentiation potential similar to existing human TSCs. Systematic omission of each, or combinations of factors, revealed the critical importance of GATA2 and GATA3 for iTSC transdifferentiation. Altogether, these findings provide insights into the transcription factor network that may be operational in the human TE and broaden the methods for establishing cellular models of early human placental progenitor cells, which may be useful in the future to model placental-associated diseases. Summary statementTranscriptional analysis of human blastocysts reveals transcription factors sufficient to derive induced trophoblast stem cells from primed human embryonic stem cells.

developmental biology↗

bioRxiv2020

KLF17 promotes human naïve pluripotency but is not required for its establishment

Current knowledge of the transcriptional regulation of human pluripotency is incomplete, with lack of inter-species conservation observed. Single-cell transcriptomics of human embryos previously enabled us to identify transcription factors, including the zinc-finger protein KLF17, that are enriched in the human epiblast and naive hESCs. Here we show that KLF17 is expressed coincident with the known pluripotency factors NANOG and SOX2 across human blastocyst development. We investigate the function of KLF17 in pluripotency using primed and naive hESCs for gain- and loss-of-function analyses. We find that ectopic expression of KLF17 in primed hESCs is sufficient to induce a naive-like transcriptome and that KLF17 can drive transgene-mediated resetting to naive pluripotency. This implies a role for KLF17 in establishing naive pluripotency. However, CRISPR-Cas9-mediated knockout studies reveal that KLF17 is not required for naive pluripotency acquisition in vitro. Transcriptome analysis of naive hESCs identifies subtle effects on metabolism and signalling following KLF17 loss of function, and possible redundancy with the related factor, KLF5. Overall, we show that KLF17 is sufficient, but not necessary, for naive pluripotency under the given in vitro conditions. Summary statementInvestigating KLF17 in human pluripotency reveals that it is sufficient, but not necessary, to establish naive hESCs. We posit that KLF17 is a peripheral regulator, like KLF2 in the mouse.

developmental biology↗

bioRxiv2020

GIANI: open-source software for automated analysis of 3D microscopy images

The study of cellular and developmental processes in physiologically relevant three-dimensional (3D) systems facilitates an understanding of mechanisms underlying cell fate, disease and injury. While cutting-edge microscopy technologies permit the routine acquisition of 3D datasets, there is currently a limited number of open-source software packages to analyse such images. Here we describe GIANI (djpbarry.github.io/Giani), new software for the analysis of 3D images, implemented as a plugin for the popular FIJI platform. The design primarily facilitates segmentation of nuclei and cells, followed by quantification of morphology and protein expression. GIANI enables routine and reproducible batch-processing of large numbers of images and also comes with scripting and command line tools, allowing users to incorporate its functionality into their own scripts and also run GIANI on a high-performance computing cluster. We demonstrate the utility of GIANI by quantifying cell morphology and protein expression in confocal images of mouse early embryos and by segmenting nuclei from light sheet microscopy images of the flour beetle embryo. We also validate the performance of the software using simulated data. More generally, we anticipate that GIANI will be a useful tool for researchers in a variety of biomedical fields.

bioinformatics↗