Biology subjects

Ku, W. L.

Publications and source records attributed to Ku, W. L..

4 recordsLinked to original sources

bioRxiv2026

Destabilization of intratumor Tregs by CTLA-4 engagement confers anti-CTLA-4-driven immunotherapy

Immune checkpoint inhibitors (ICI) that are antibodies against CTLA-4 have achieved therapeutic effects on multiple types of cancers 1-4, but the mechanisms underlying the therapy remain incompletely understood. In contrast to the initial theory that the antibody blockades CTLA-4 on T cells is of central importance, it has recently been demonstrated that selective reduction of CD4+Foxp3+ regulatory T cells (Tregs) in the tumor microenvironment by the antibody plays a key role in the anti-tumor effects5-8, although this phenomenon remains under debate in human patients9-12. We show here that anti-CTLA-4 antibody engages CTLA-4 in Tregs specifically in the tumor tissues to reduce their stability and survival and weaken their suppressive function through upregulating TGF-{beta} signaling. This leads to the antibody-mediated - upregulation of CD4+ and CD8+ effector T cell anti-tumor immunity and consequently cancer immunotherapy. Specifically, anti-CTLA-4 antibody directly stimulates CTLA-4 in intratumor Tregs to enhance their TGF-{beta} receptor I (T{beta}RI)-mediated TGF-{beta} signaling13. This results in reduction of IL-2 receptor CD25 expression and downregulation of lactate metabolism in intratumor Tregs to reduce their stability and survival 14-16, and also compromises their suppressive function by inhibiting Foxp3 expression. This activity requires high levels of CTLA-4 expression in the intratumor Tregs and the presence of antibody Fc receptors in the tumor tissues. Strikingly, deletion of T{beta}RI specifically in Tregs completely prevents the reduction of intratumor Tregs and abrogates the anti-CTLA-4-mediated cancer immunotherapy. In contrast to intratumor Tregs, anti-CTLA-4 antibody treatment blocks CTLA-4 in the intratumor CD4+ Foxp3- and CD8+ T effector cells and in the peripheral Tregs to decrease their T{beta}RI expression and increase their expansion due to their relatively low levels of CTLA-4 in the same tumor bearing mice. Significantly, the engagement of CTLA-4 by anti-human CTLA-4 antibody (ipilimumab) also upregulates TGFBR1 but decreases Il2RA expression and lactate metabolism in human Tregs in vitro and in humanized mice in vivo, leading to suppression of tumor progression. We have provided additional mechanism underlying anti-CTLA-4-mediated anti-tumor effects through destabilizing intratumor Tregs by CTLA-4 engagement-mediated TGF-{beta} signaling. This could lay a theoretical foundation for designing optimal immunotherapy based on anti-CTLA-4 antibody in cancer patients.

immunology↗

bioRxiv2024

The Drosophila histone methyl-transferase SET1 coordinates multiple signaling pathways in regulating male germline stem cell maintenance and differentiation

Many cell types come from tissue-specific adult stem cells that maintain the balance between proliferation and differentiation. Here, we study how the H3K4me3 methyltransferase, Set1, regulates early-stage male germ cell proliferation and differentiation in Drosophila. Early-stage germline-specific knockdown of set1 results in a temporally progressed defects, arising as germ cell loss and developing to overpopulated early-stage germ cells. These germline defects also impact the niche architecture and cyst stem cell lineage in a non-cell-autonomous manner. Additionally, wild-type Set1, but not the catalytically inactive Set1, could rescue the set1 knockdown phenotypes, highlighting the functional importance of the methyl-transferase activity of the Set1 enzyme. Further, RNA-seq experiments reveal key signaling pathway components, such as the JAK-STAT pathway gene stat92E and the BMP pathway gene mad, that are upregulated upon set1 knockdown. Genetic interaction assays support the functional relationships between set1 and JAK-STAT or BMP pathways, as mutations of both the stat92E and mad genes suppress the set1 knockdown phenotypes. These findings enhance our understanding of the balance between proliferation and differentiation in an adult stem cell lineage. The germ cell loss followed by over-proliferation phenotypes when inhibiting a histone methyl-transferase raise concerns about using their inhibitors in cancer therapy.

developmental biology↗

bioRxiv2022

Roles of the distinct N-terminal amino acid between H3 and H3.3 in Drosophila male germline stem cell lineage

Adult stem cells undergo asymmetric cell divisions to produce two daughter cells with distinct cell fates: one capable of self-renewal and the other committed for differentiation. Mis-regulation of this delicate balance can lead to cancer and tissue degeneration. During asymmetric division of Drosophila male germline stem cells (GSCs), preexisting (old) and newly synthesized histone H3 are differentially segregated whereas old and new histone variant H3.3 are more equally inherited. However, what underlies these distinct inheritance patterns remains unknown. Here, we report that the N-terminal tails of H3 and H3.3 are critical for their inheritance patterns, as well as GSC maintenance and proper differentiation. H3 and H3.3 differ at the 31st position in their N-termini with Alanine for H3 and Serine for H3.3. By swapping these two amino acids, we generated two mutant histones (i.e., H3A31S and H3.3S31A). Upon expressing them in the early-stage germline, we identified opposing phenotypes: over-population of early-stage germ cells in the H3A31S-expressing testes and significant germ cell loss in testes expressing the H3.3S31A. Asymmetric H3 inheritance is disrupted in the H3A31S-expressing GSCs, due to mis-incorporation of old histones between sister chromatids during DNA replication. Furthermore, H3.3S31A mutation accelerates old histone turn-over in the GSCs. Finally, using a modified Chromatin Immunocleavage assay on early-stage germ cells, we found that H3A31S has an enhanced occupancy at the promoters and transcription starting sites than H3, while H3.3S31A is more enriched at transcriptionally silent intergenic regions compared to H3.3. Overall, these results suggest that the 31st amino acids for both H3 and H3.3 are critical for their proper genomic occupancy and function. Together, our findings indicate a critical role for the different amino acid composition of the N-terminal tails between H3 and H3.3 in an endogenous stem cell lineage, and provide insights into the importance of proper histone inheritance in specifying cell fates and regulating cellular differentiation.

developmental biology↗

bioRxiv2019

Mapping Histone Modifications in Low Cell Number and Single Cells Using Antibody-guided Chromatin Tagmentation (ACT-seq)

Modern next-generation sequencing-based methods have empowered researchers to assay the epigenetic states of individual cells. Existing techniques for profiling epigenetic marks in single cells often require the use and optimization of time-intensive procedures such as drop fluidics, chromatin fragmentation, and end repair. Here we describe ACT-seq, a novel and streamlined method for mapping genome-wide distributions of histone tail modifications, histone variants, and chromatin-binding proteins in a small number of or single cells. ACT-seq utilizes a fusion of Tn5 transposase to Protein A that is targeted to chromatin by a specific antibody, allowing chromatin fragmentation and sequence tag insertion specifically at genomic sites presenting the relevant antigen. The Tn5 transposase enables the use of an index multiplexing strategy (iACT-seq), which enables construction of thousands of single-cell libraries in one day by a single researcher without the need for drop-based fluidics or visual sorting. We conclude that ACT-seq present an attractive alternative to existing techniques for mapping epigenetic marks in single cells.

genomics↗