Biology subjects

Pae, J.

Publications and source records attributed to Pae, J..

7 recordsLinked to original sources

bioRxiv2026

Clonal dynamics of germinal center refueling by secondary immunization

Many vaccine regimens involve delivery of multiple doses to the same anatomical site, such that booster doses frequently encounter germinal centers (GCs) still active from prior immunization. The consequences of this "GC refueling" to B cell clonality have not been systematically investigated. Using mouse models of mRNA-LNP vaccination combined with multicolor fate-mapping, longitudinal GC imaging, and immunoglobulin sequencing, we show that refueling triggers clonal burst-type expansion of GC-resident B cells, rather than recruiting local memory, resulting in marked focusing of GCs on the descendants of individual B cells. Refueling with a drifted antigen led to limited but detectable retraining of primary-cohort clones, although most variant-specific responses in this setting arose from newly recruited naive B cells. These findings identify GC refueling as a distinct mode of vaccine response with implications for sequential immunization strategies against rapidly evolving pathogens.

immunology↗

bioRxiv2025

GCL pruning of PIP3 establishes the soma-germline boundary

Primordial germ cells (PGCs) are the first cells specified in the Drosophila embryo and serve as precursors to the germline. Their formation requires suppression of somatic fates, a process achieved by excluding the receptor tyrosine kinase Torso from the posterior pole through degradation mediated by the ubiquitin ligase adaptor Germ Cell-Less (GCL). Although Torso is known to antagonize PGC formation, the underlying mechanism has remained unclear. Here, we combine optogenetic Ras activation and Ras effector loop mutants to show that Ras signaling suppresses PGC formation independently of the canonical Raf/MEK/ERK pathway. We identify an unexpected early role for Torso in activating phosphoinositide 3-kinase (PI3K), generating posterior membrane domains enriched in phosphatidylinositol (3,4,5)-trisphosphate (PIP3). Elevated PI3K activity disrupts PGC formation, while reduced PI3K activity leads to ectopic PGCs. We further demonstrate that GCL remodels the posterior pole membrane by suppressing Torso-dependent PI3K activation. Clearing PIP3 enables Myosin II enrichment, thereby constricting the pole bud for PGC formation. Together, our findings reveal how antagonistic Torso and GCL activities establish the soma-germline boundary by regulating cortical lipid organization. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/697122v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@1efe138org.highwire.dtl.DTLVardef@1ab3b17org.highwire.dtl.DTLVardef@b49412org.highwire.dtl.DTLVardef@1655c5e_HPS_FORMAT_FIGEXP M_FIG Graphical AbstractO_LIMembrane domains with high Torso activity (purple) in the early embryo have a higher PIP3 content. C_LIO_LIAt the posterior pole, GCL-containing germplasm (green) degrades Torso, resulting in a PIP3-low membrane. C_LIO_LIClearing PIP3 enables Myosin II pole bud constriction required for PGC formation. C_LI C_FIG

cell biology↗

bioRxiv2025

Direct measurement of sub-kilobase chromatin structure reveals that linker histone H1 broadly compacts chromatin, with differential impact amongst epigenetic states

Chromatin compaction by linker histone H1 family proteins is a long-standing model for transcriptional repression. However, the biophysical and conformational details of such compaction in situ, at the kilobase- and sub-kilobase length scale relevant to the activity of transcriptional regulatory elements, remain under debate. Rather than inferring such compaction from indirect measurements of features like DNA accessibility, we sought to directly probe sub-kilobase contacts between nearby nucleosomes. We developed an improved version of radiation-induced correlated cleavage with sequencing (RICC-seq), which we term RICC-seq 2.0, and used it in parallel with Micro-C to cross-validate our measurements of chromatin structure in both diverse cell types with different levels of linker histone and different levels of chromatin compaction, as well as a CRISPRi system for pan-H1 depletion. Using this system, we find that chromatin fiber de-compaction upon H1 depletion is global across the genome, reducing the contrast in inter-nucleosome contacts between acetylated chromatin and the rest of the genome. Surprisingly, this does not dramatically change higher-order chromatin organization such as nuclear compartments. Nevertheless, we observe a broad increase in accessibility at tens of thousands of sites and an increase in expression of over a thousand genes, which are enriched in polycomb repressive complex targets. Investigating the local chromatin compaction at upregulated genes as opposed to genes that do not change transcription, we observe that upregulated genes are not specifically de-compacted. Rather, our data support a model in which linker histone globally induces local compaction of nucleosome contacts and an increase in linker lengths, and repression by PRC1/2 is particularly dependent on these local features of chromatin architecture.

genomics↗

bioRxiv2025

Replaying germinal center evolution on a quantified affinity landscape

Darwinian evolution of immunoglobulin genes within germinal centers (GCs) underlies the progressive increase in antibody affinity following antigen exposure. Whereas the cellular mechanics of how competition between B cells produces increases in affinity are well established, the evolutionary dynamics of this process are less clear. We developed an experimental evolution model where we "replay" over one hundred monoclonal GC reactions, assigning affinities to each cell using deep mutational scanning. Our data reveal how GCs achieve predictable outcomes by means of noisy but persistent selection on an affinity landscape whose exploration is heavily constrained by somatic hypermutation biases. We infer a fitness landscape that quantitatively recapitulates the affinity maturation trajectory of our clone and find that apparent features of GC selection such as permissiveness to low-affinity lineages and rapid plateauing of affinity are likely artifacts of survivorship biases that distort our view of how B cell affinity progresses over time.

immunology↗

bioRxiv2025

GMCL1 Controls 53BP1 Stability and Modulates Paclitaxel Sensitivity in Cancer

Mitotic surveillance pathways monitor the duration of mitosis (M phase) in the cell cycle. Prolonged M phase, caused by spindle attachment defects or microtubule-targeting drugs triggers formation of the ternary "mitotic stopwatch pathway" complex (MSP) consisting of 53BP1, USP28, and p53. This complex stabilizes p53, leading to cell cycle arrest or apoptosis in daughter cells. In cancers that are resistant to paclitaxel, a microtubule-targeting agent, cells bypass mitotic surveillance activation, allowing unchecked proliferation, although the underlying mechanisms remain poorly understood. Here, we identify GMCL1 as a key negative regulator of MSP signaling. We show that 53BP1 physically interacts with GMCL1, but not its paralog GMCL2, and we map their interaction domains. CRL3GMCL1 functions as a ubiquitin ligase that targets 53BP1 for degradation during the M phase, thereby reducing p53 accumulation in daughter cells. Depletion of GMCL1 inhibits cell cycle progression upon release from prolonged mitotic arrest, a defect that is rescued by co-silencing 53BP1 or USP28. Moreover, GMCL1 depletion sensitizes cancer cells to paclitaxel in a p53-dependent manner. Together, our findings support a model in which dysregulated CRL3GMCL1-mediated degradation of 53BP1 prevents proper MSP function, leading to p53 degradation and continued proliferation. Targeting GMCL1 may therefore represent one possible avenue for addressing paclitaxel resistance in cancer cells with functional p53.

cell biology↗

bioRxiv2022

Proximity-dependent labeling identifies dendritic cells that prime the antitumor CD4+ T cell response

Dendritic cells (DCs) are uniquely capable of transporting tumoral antigens to tumor-draining lymph nodes (tdLNs), and also interact with effector T cells within the tumor microenvironment (TME) itself, mediating both natural antitumor immunity and the response to checkpoint blockade immunotherapy. Using LIPSTIC (Labeling Immune Partnerships by SorTagging Intercellular Contacts)-based single-cell transcriptomics, we identify individual DCs capable of presenting antigen to CD4+ T cells in the tdLN as well as inside the tumor microenvironment (TME). Our findings reveal that DCs with similar hyperactivated transcriptional phenotypes interact with helper T cells both within tumors and in the tdLN, and that checkpoint blockade drugs enhance these interactions. These findings show that a relatively small fraction of DCs is responsible for most of the antigen presentation within the tdLN and TME to both CD4+ and CD8+ tumor-specific T cells and that classical checkpoint blockade enhances CD40-driven DC activation at both sites.

immunology↗

bioRxiv2020

Cyclin D3 drives inertial cell cycling in dark zone germinal center B cells

During affinity maturation, germinal center (GC) B cells alternate between proliferation and so-matic hypermutation in the dark zone (DZ) and affinity-dependent selection in the light zone (LZ). This anatomical segregation imposes that the vigorous proliferation that allows clonal expansion of positively-selected GC B cells takes place ostensibly in the absence of the signals that triggered selection in the LZ, as if by "inertia." We find that such inertial cycles specifically require the cell cycle regulator cyclin D3. Cyclin D3 dose-dependently controls the extent to which B cells proliferate in the DZ and is essential for effective clonal expansion of GC B cells in response to strong T follicular helper (Tfh) cell help. Introduction into the Ccnd3 gene of a Burkitt lymphoma-associated gain-of-function mutation (T283A) leads to larger GCs with increased DZ proliferation and, in older mice, to clonal B cell lymphoproliferation, suggesting that the DZ inertial cell cycle program can be coopted by B cells undergoing malignant transformation.

immunology↗