Biology subjects

Khalighifar, A.

Publications and source records attributed to Khalighifar, A..

3 recordsLinked to original sources

bioRxiv2026

Shaping cryogenic 3D volume imaging by pFIB ion species and milling voltage

Cryogenic plasma focused ion beam-scanning electron microscopy (cryo-pFIB-SEM) enables nanoscale volume imaging of vitrified cells and tissues, but the fidelity of each newly exposed block-face depends on how ions interact with compositionally heterogeneous biological material. Using xenon, argon, oxygen, and nitrogen plasma beams across a 30-2 kV voltage range, we examined how ion species and accelerating voltage influence block-face quality and ultrastructural fidelity. Curtaining imposed a species-dependent lower limit on accelerating voltage, with xenon and argon supporting uniform milling to 10 kV and oxygen and nitrogen to 15 kV. Monte Carlo simulations predicted substantially reduced ion penetration and atomic displacement under these species-specific low-kV conditions. Experimentally, lowering milling voltage suppressed charging and topographic artifacts, with the largest effects observed at heterogeneous interfaces. Oxygen and nitrogen produced greater membrane sharpness and organelle contrast than the noble gases, and these differences influenced machine-learning-based recognition of fine organelle features in 3D datasets. Together, these findings show that cryo-pFIB-SEM image quality cannot be optimized by accelerating voltage or ion species independently. Instead, milling performance reflects a tradeoff among surface fidelity, charging, contrast, and curtaining that depends on local specimen composition. Species-specific low-kV milling therefore provides a practical framework for matching cryo-pFIB-SEM acquisition conditions to the structural and biophysical properties of the biological target.

cell biology↗

bioRxiv2026

Protein domain characterization reveals human MIC60 tolerates loss of helical bundle domain

The mitochondrial contact site and cristae organizing system (MICOS) is essential for cristae junction formation and inner mitochondrial membrane architecture. To define how MICOS integrity is established and maintained, we generated conditional deletion models of Immt (encoding MIC60), a core MICOS subunit, in tissue-specific settings and in cultured cells. Liver-specific deletion of Immt in mice induced profound defects in mitochondrial ultrastructure and function, establishing MIC60 as essential for mitochondrial integrity. Notably, despite the severity of the defects, we did not detect increased apoptosis in liver tissue or in cells. To directly link MIC60 structure to its function, we performed a systemic structure-function analysis of human MIC60 using domain-specific deletion mutants expressed in Immt-deleted cells. We identified that the transmembrane, coiled-coil, and mitofilin domains are required for MICOS assembly, mitochondrial morphology, and respiratory function. Unexpectedly, deletion of the predicted helical bundle (a region spanning 229 amino acids) substantially restored mitochondrial structure and function, nearly matching full-length MIC60. A mutation (K299E) associated with human disease within this domain similarly preserved most MIC60-dependent functions. Together, these results establish MIC60 as a non-redundant regulator of mitochondrial architecture while revealing that a large predicted structural domain is largely dispensable for MIC60s core functions, refining current models of MICOS organization and uncovering unexpected modularity within MIC60.

cell biology↗

bioRxiv2024

MOZ and HBO1 Histone Acetyltransferase Complexes Are Molecular Dependencies and Therapeutic Targets in NUP98-Rearranged Acute Myeloid Leukemia

NUP98 fusion oncoproteins (FOs) are a hallmark of childhood acute myeloid leukemia (AML) and drive leukemogenesis through liquid-liquid phase separation-mediated nuclear condensate formation. However, the composition and consequences of NUP98 FO-associated condensates are incompletely understood. Here we show that MYST family histone acetyltransferase (HAT) complex proteins including MOZ/KAT6A, HBO1/KAT7, and the common MOZ/HBO1 complex subunit BRPF1 associate with NUP98 FOs on chromatin and within condensates. MYST HATs are molecular dependencies in NUP98-rearranged (NUP98-r) leukemia, and genetic inactivation or pharmacologic inhibition of Moz and Hbo1 impairs NUP98-r cell fitness. MOZ/HBO1 inhibition decreased global H3K23ac levels, displaced NUP98::HOXA9 from chromatin at the Meis1 locus, and led to myeloid cell differentiation. Additionally, MOZ/HBO1 inhibition decreased leukemic burden in multiple NUP98-r leukemia xenograft mouse models, synergized with Menin inhibitor treatment, and was efficacious in Menin inhibitor-resistant cells. In summary, we show that MYST family HATs are therapeutically actionable dependencies in NUP98-r AML. SIGNIFICANCE STATEMENTMOZ and HBO1 associate with NUP98 fusion oncoprotein condensates to drive leukemogenesis. Inhibition of their histone acetyltransferase activity is an effective therapeutic strategy in NUP98-rearranged leukemias, including those resistant to Menin inhibition. Moreover, combined MOZ/HBO1 and Menin inhibition is synergistic, supporting clinical translation to improve outcomes of NUP98 FO-driven leukemias.

cancer biology↗