Biology subjects

Hsieh, S.-C.

Publications and source records attributed to Hsieh, S.-C..

6 recordsLinked to original sources

bioRxiv2026

Rhizobia transform legume nodules via native T-DNA transfer systems

Agrobacteria can deliver transfer DNA (T-DNA) into plant cells, causing tumorous growth. This process, agrobacteria-mediated transformation (AMT), underlies plant genetic engineering and has been regarded as an evolutionary innovation specific to these pathogens. By screening 63,759 alphaproteobacterial genomes for T-DNA border sequences, we identify mini-T-DNAs in diverse rhizobial genera. Each typically encodes a single cargo, a RolB/RolC family protein or a distant rhizobial homologue. We show that native rhizobia-mediated transformation (RMT) systems deliver these mini-T-DNAs into the root nodules of common bean, pea, and chickpea, where the cargo genes are expressed, with no signal in adjacent root tissue. RMT systems share VirG-dependent activation of the core virulence regulon but respond to distinct upstream signals. Our genome screen identifies many additional candidate T-DNA transfer systems, and phylogenetic analysis places their genes into two of the many type IV secretion system (T4SS) clades. RMT systems occur in both clades, together with other systems unrelated to T-DNA transfer, and typically have an ancestral-like T4SS configuration. In contrast, AMT systems occupy a single subclade, consistent with later acquisition of their T4SS module. AMT and RMT thus represent two specialized outcomes of an older, more widespread capability for bacteria-to-plant DNA transfer.

microbiology↗

bioRxiv2025

Tn7 family bacterial transposons known for tightly controlled target site selection are distributed across Asgard and archaeal groups

Tn7 family elements are DNA transposons in bacteria that show tight control of targeting, including multiple varieties that evolved guide RNA-directed transposition. We find diverse Tn7 elements across all archaeal superphyla, including many examples in the Asgard group where the branchpoint with eukaryotes is believed to reside. Mirroring what is known from Tn7 elements in bacteria, specialized specific sites were targeted like tRNA genes and housekeeping genes like ribonucleotide reductase. Reconstituting an Asgard element confirmed that the element possesses the hallmark properties of the Tn7 family. Tn7 elements in bacteria evolve new targeting pathways by recursive cooption of DNA binding domains a process that was also found with protein domains from the eukaryote-archaea sisterhood in the archaeal elements. This work expands our understanding of mobile DNA diversity in archaea and the role a specialized DNA transposon family plays in gene transfer within and between domains.

microbiology↗

bioRxiv2024

Telomeric transposons are pervasive in linear bacterial genomes

Eukaryotes have linear DNA and their telomeres are hotspots for transposons, which in some cases took over telomere maintenance. While many bacteria also have linear chromosomes and plasmids, no transposons were known to target bacterial telomeres. Here we show several families of independently evolved telomeric transposons in cyanobacteria and Streptomyces. While these elements have one specific transposon end sequence with the second boundary being the telomere, we can show they move using two transposon ends. Telomeres are transiently bridged by the telomere maintenance systems, providing a duplex substrate for mobilization of the element and the associated telomere. We identify multiple instances where telomeric transposons have replaced native telomeres, making the host cell dependent on the new telomere system for genome maintenance. This work indicates how telomeric transposons can promote gene transfer both between and within genomes, significantly influencing the evolutionary dynamics of linear genomes.

microbiology↗

bioRxiv2021

Structural basis for target-site selection in RNA-guided DNA transposition systems

CRISPR-associated transposition systems allow guide RNA-directed integration of a single DNA insertion in one orientation at a fixed distance from a programmable target sequence. We define the mechanism explaining this process by characterizing the transposition regulator, TnsC, from a Type V-K CRISPR-transposase system using cryo-EM. Polymerization of ATP-bound TnsC helical filaments explains how polarity information is passed to the transposase. Our Cryo-EM structure of TniQ-TnsC reveals that TniQ caps the TnsC filament, establishing a universal mechanism for target information transfer in Tn7/Tn7-like elements. Transposase-driven disassembly establishes delivery of the element only to unused protospacers. Finally, structures with the transition state mimic, ADP{middle dot}AlF3, reveals how TnsC transitions to define the fixed point of insertion. These mechanistic findings provide the underpinnings for engineering CRISPR-associated transposition systems for research and therapeutic applications. One Sentence SummaryCryo-EM studies reveals the role of the AAA+ regulator TnsC for target-site selection in CRISPR-associated transposition systems.

molecular biology↗

bioRxiv2021

Tn7-CRISPR-Cas12K elements manage pathway choice using truncated repeat-spacer units to target tRNA attachment sites

CRISPR-Cas systems provide a defense against mobile elements. These defense systems have been naturally coopted multiple times for guide RNA-directed transposition by Tn7-like transposons. Elements associated with a type I-F CRISPR-Cas system categorize guide RNAs, maintaining a standard CRISPR array capable of acquiring new spacers targeting other mobile elements while maintaining a special guide RNA allowing integration into a conserved site in the chromosome called an attachment site. We show here that Tn7-like elements associated with a type V-K (Cas12K-based) system use a similar strategy to target diverse tRNA genes as attachment sites. These guides are encoded as truncated minimal repeat-spacer units and are found in distinct locations. Multiple pieces of information support that V-K guide RNAs are acquired using a type I-D adaptation system, but remain private to the V-K transposition process. This catalog of Cas12K elements and naturally occurring insertions will help future work engineering precision integration systems.

molecular biology↗

bioRxiv2020

Guide RNA categorization enables target site choice in Tn7-CRISPR-Cas transposons

SummaryCRISPR-Cas defense systems have been coopted multiple times in nature for guide RNA-directed transposition by Tn7-like elements. Prototypic Tn7 uses dedicated proteins for two targeting pathways, one targeting a neutral and conserved attachment site in the chromosome and a second directing transposition into mobile plasmids facilitating cell-to-cell transfer. We show that Tn7-CRISPR-Cas elements evolved a system of guide RNA categorization to accomplish the same two-pathway lifestyle. Selective regulation of specialized guide RNAs allows long-term memory for access to chromosomal sites upon entry into a new host, while conventional CRISPR features maintain the ability to continually acquire guide RNAs to new plasmid and phage targets. Transposon-encoded guide RNAs are also privatized to be recognized only by the transposon-adapted system working with selective regulation to guard against toxic self-targeting by endogenous CRISPR-Cas defense systems. This information reveals new avenues to engineer guide RNAs for enhanced CRISPR-Cas functionality for genome modification.Competing Interest StatementCornell University has filed patent applications related to this work with JEP as inventor.View Full Text

molecular biology↗