Biology subjects

Huo, T.

Publications and source records attributed to Huo, T..

4 recordsLinked to original sources

bioRxiv2024

The assembly and efflux processes revealed by in situ structures of the ABC-type tripartite pump MacAB-TolC

The MacAB-TolC tripartite efflux pump is widely distributed in Gram-negative bacteria, extruding antibiotics and virulence factors that lead to multidrug resistance and pathogenicity. This pump spans the cell envelope through its three components, an inner membrane ATP-binding cassette (ABC) transporter MacB, a periplasmic adaptor protein MacA, and an outer membrane protein TolC. However, the assembly process and efflux mechanism of the MacAB-TolC in cells remain unclear. Here, we resolve the in situ structures of Escherichia coli (E. coli) MacAB-TolC efflux pump by electron cryo-tomography and subtomogram averaging. In E. coli without antibiotic treatment, we observe a fully assembled MacAB-TolC pump with a weak constriction density in the middle of the MacA region. When E. coli cells were treated with erythromycin, we discovered the emergence of MacA-TolC subcomplexes, indicating flexible binding of MacB in the presence of an antibiotic substrate. This finding was further validated by in vivo crosslinking results. Together, our data present the in situ assembly process of the MacAB-TolC and derive a substrate-driven working model for ABC-type tripartite pump.

microbiology↗

bioRxiv2023

In situ structures of secretins from bacterial type II secretion system reveal their membrane interactions and translocation process

The GspD secretin is the outer membrane channel of the bacterial type II secretion system (T2SS) which secrets diverse effector proteins or toxins that cause severe diseases such as diarrhea and cholera. GspD needs to translocate from the inner to the outer membrane to exert its function, and this process is an essential step for T2SS to assemble. Here, we investigate two types of secretins discovered so far in Escherichia coli, GspD and GspD{beta}, respectively. By electron cryotomography subtomogram averaging, we determine in situ structures of all the key intermediate states of GspD and GspD{beta} in the translocation process, with resolution ranging from 9 [A] to 19 [A]. In our results, GspD and GspD{beta} present entirely different membrane interaction patterns and ways of going across the peptidoglycan layer. We propose two distinct models for the membrane translocation of GspD and GspD{beta}, providing a comprehensive perspective on the inner to outer membrane biogenesis of T2SS secretins.

microbiology↗

bioRxiv2022

Full-length αIIbβ3 CryoEM structure reveals intact integrin initiate-activation intrinsic architecture

Integrin IIb{beta}3 is the key receptor regulating platelet retraction and accumulation, thus pivotal for hemostasis, and arterial thrombosis as well as a proven drug-target for antithrombotic therapies. Here we resolve the cryoEM structures of the intact full-length IIb{beta}3, which covers three distinct states along the activation pathway. Here, we resolve intact IIb{beta}3 structure at 3[A] resolution, revealing the overall topology of the heterodimer with the transmembrane (TM) helices and the head region ligand-binding domain tucked in a specific angle proximity to the TM region. In response to the addition of a Mn2+ agonist, we resolved two coexisting states, "intermediate" and "pre-active". Our structures show conformational changes of the intact IIb{beta}3 activating trajectory, as well as a unique twisting of the lower integrin legs representing an intermediate state (TM region at a twisting conformation) and a coexisting pre-active state (bent and opening in leg), which is required for inducing the transitioning platelets to accumulate. Our structure provides for the first time direct structural evidence for the lower legs involvement in full-length integrin activation mechanisms. Additionally, our structure offers a new strategy to target the IIb{beta}3 lower leg allosterically instead of modulating the affinity of the IIb{beta}3 head region.

biochemistry↗

bioRxiv2021

Cryo-ET distinguishes platelets during pre-acute myeloid leukemia from steady state hematopoiesis

Early diagnosis of acute myeloid leukemia (AML) in the pre-leukemic stage remains a clinical challenge, as pre-leukemic patients show no symptoms, lacking any known morphological or numerical abnormalities in blood cells. Here, we demonstrate that platelets with structurally abnormal mitochondria emerge at the pre-leukemic phase of AML, preceding detectable changes in blood cell counts or detection of leukemic blasts in blood. We visualized frozen-hydrated platelets from mice at different time points during AML development in situ using electron cryo-tomography (cryo-ET) and identified intracellular organelles through an unbiased semi-automatic process followed by quantitative measurement. A large proportion of platelets exhibited changes in the overall shape and depletion of organelles in AML. Notably, 23% of platelets in pre-leukemic cells exhibit abnormal, round mitochondria with unfolded cristae, accompanied by a significant drop in ATP levels and altered expression of metabolism-related gene signatures. Our study demonstrates that detectable structural changes in pre-leukemic platelets may serve as a biomarker for the early diagnosis of AML.

biophysics↗