Biology subjects

Hahn, S.

Publications and source records attributed to Hahn, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Loss of Mitochondrial Respiratory Capacity Reshapes Myeloid Cell Function during Mycobacterium tuberculosis infection

Myeloid cells are essential mediators of host defense against Mycobacterium tuberculosis (Mtb), yet the metabolic programs that sustain their function during chronic infection remain poorly defined. Here, using scRNA-seq we identified a striking, coordinated decline in mitochondrial electron transport chain gene expression across diverse myeloid populations as Mtb disease progressed in mice. This transcriptional remodeling was associated with broad changes in immune and metabolic pathways, including reduced antigen presentation, interferon responses, protein synthesis, and glycolysis. Accordingly, loss of Complex I in macrophages (Ndufs4 knockdown) reduced MHC-II surface expression, dysregulated inflammatory gene expression, and limited control of Mtb replication. Finally, analysis of single-cell transcriptomic data from Mtb-exposed human household contacts identified an almost identical transcriptional program enriched in IGRA+ individuals, supporting a role for mitochondrial respiratory remodeling in human TB. Together, these findings demonstrate that mitochondrial bioenergetic competence is required to sustain macrophage effector function during chronic Mtb infection and suggest that mitochondrial restoration may boost protective responses in TB patients.

immunology↗

bioRxiv2026

Energetic benefits of differential migration strategies shift under climate change for a migratory bird

Understanding how migratory birds balance energetic costs of movement with wintering benefits requires quantifying their energy expenditure across the full annual cycle. Here, we present a novel approach to reconstruct annual energy budget from multi-sensor geolocator data on atmospheric pressure and activity. We apply this method to tracking data of Eurasian Wrynecks from a European breeding population exhibiting striking variation in migration distance: short-distance migration to southern Europe, medium-distance to northern Africa, and long-distance to sub-Saharan Africa. Long-distance migrants had 21-26% lower annual energy expenditure than shorter-distance migrants, primarily due to reduced thermoregulation costs during boreal winter. However, they faced extreme physiological demands during migration, with daily energy expenditure exceeding 9.5 times basal metabolic rate. Since 1950, climate warming has progressively reduced winter thermoregulation demands disproportionately benefiting and potentially promoting shorter-distance strategies. These results reveal shifting energetic trade-offs under climate change, potentially driving evolution of migration patterns.

ecology↗

bioRxiv2025

HnRNPA2B1 tunes antimycobacterial immune responses in macrophages through alternative splicing of Irgm1

Onset and progression of active tuberculosis disease result from upsetting the delicate balance between Mtb virulence and host defenses. Because it dynamically tunes the functional output of protein expression in cells, alternative splicing, a process by which different mRNAs can be gen-erated from a single gene, is positioned to play a critical role in maintaining an equilibrated Mtb-macrophage host-pathogen interface. To gain insight into how alternative splicing shapes anti-mycobacterial immune responses, we used RNA-sequencing and splicing-aware computational pipelines to quantify alternative splicing in Mtb-infected bone marrow-derived murine macro-phages. We found that [~]5% of expressed macrophage genes exhibit one or more splicing changes at 8h post-Mtb infection, highlighting alternative splicing as a key regulatory node in the macrophage response to Mtb. We next sought to identify RNA binding proteins that play an out-sized role in shaping the macrophage transcriptome during Mtb infection. We discovered that the splicing factor heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1) promotes the early induction of inflammatory genes while dampening several type I interferon-stimulated genes in response to Mtb. HnRNPA2B1 also controls alternative splicing of many genes during Mtb infection, including Irgm1, a critical immunity-related GTPase. The balance of Irgm1-long vs. -short is differentially regulated in response to diverse inflammatory cues and macrophages overexpressing Irgm1-short are defective in autophagosomal targeting, lysosomal homeostasis, and restriction of Mtb replication. These data highlight a key role for AS in shaping the macro-phage transcriptome and pinpoint hnRNPA2B1 as a novel restriction factor in the cell-intrinsic response to Mtb. IMPORTANCEAlthough the process of making proteins from RNAs requires many steps (transcription, cap-ping/polyadenylation, pre-mRNA splicing, mRNA export, mRNA modifications, etc.), we know very little about how post-transcriptional steps contribute to host immune defenses. Here, we show that alternative splicing, the process of making different mature RNAs from a single pre-cursor RNA, is a prominent and dynamic feature of macrophage infection with the bacterial pathogen Mycobacterium tuberculosis (Mtb). We identify the splicing regulator hnRNPA2B1 as a key coordinator of early gene expression during Mtb infection, influencing pathways that pro-mote inflammation and help restrict bacterial growth. Notably, we report that hnRNPA2B1 con-trols the splicing of the antimycobacterial protein Irgm1 to generate different flavors of the pro-tein. Since only one Irgm1 flavor can restrict Mtb growth inside macrophages, maintaining the balance of these proteins in response to diverse inflammatory cues is important. By revealing how RNA processing shapes the macrophage response to Mtb, our work highlights an often-overlooked layer of immune regulation and opens new avenues for splicing-targeted therapies designed to boost Mtb killing in macrophages.

microbiology↗

bioRxiv2025

YAP/TEAD4-reagulated placental alkaline phosphatases ALPP and ALPPL2 are immunosuppressive ectonucleotidases modulated by MAPK inhibitors

Placental alkaline phosphatases ALPP and ALPPL2 (ALPP/L2) are promising clinical targets for therapies such as antibody-drug conjugates (ADCs). However, their regulation and biological functions remain unclear. Here, we identify the YAP/TEAD4 signaling axis as the primary regulator of ALPP/L2 expression in tumor cells. Importantly, we demonstrate that inhibitors targeting KRAS, MEK, and ERK upregulate ALPP/L2 expression in vitro and in vivo, thereby enhancing the efficacy of antibody-drug conjugates (ADCs) when used in combination. Moreover, our work reveals a novel immunomodulatory function of ALPP/L2 as cell surface ectonucleotidases that hydrolyze extracellular ATP, ADP, and AMP to adenosine, promoting regulatory T cell infiltration into the tumor microenvironment. Together, these findings reveal a biological function of ALPP/L2 and identify druggable regulators to enhance ALPP/L2-targeted therapies. SIGNIFICANCEThis study uncovers the YAP/TEAD pathway as a positive regulator of the clinically relevant tumor-associated antigens ALPP and ALPPL2. Kras, MEK, or ERK inhibitors enhance ALPP/L2 expression in tumor cells, thereby increasing the efficacy of ALPP/L2-targeted antibody-drug conjugates. These findings offer a promising combination strategy to overcome resistance to ALPP/L2-based therapies caused by low target expression.

cancer biology↗

bioRxiv2024

Transcriptional bursting, gene activation, and roles of SAGA and Mediator Tail measured using nucleotide recoding single cell RNA-seq

A time resolved nascent single-cell RNA-seq approach was developed to dissect gene-specific transcriptional bursting and the roles of SAGA and Mediator Tail (the activator-binding module). Most yeast genes show near-constitutive behavior while only a subset of genes show high mRNA variance suggestive of transcription bursting. Bursting behavior is highest in the coactivator redundant (CR) gene class (dependent on both SAGA and TFIID) and is strongest in TATA-containing CR genes. Applying this approach to analyze gene activation, we found that basal histone gene transcription is in a low level, low-noise constitutive mode while the activated state unexpectedly shows an increase in both the fraction of active promoters and a switch to a noisy and bursty transcription mode. Rapid depletion of either SAGA or Mediator Tail suggests that both factors play an important role in stimulating the fraction of active promoters at CR genes, with a variable gene-specific role in transcriptional bursting.

genomics↗

bioRxiv2022

Broad compatibility between yeast UAS elements and core promoters and identification of promoter elements that determine cofactor specificity

Three general classes of yeast protein-coding genes are distinguished by their dependence on the transcription cofactors TFIID, SAGA and Mediator (MED) Tail, but little is known about whether this dependence is determined by the core promoter, Upstream activation sites (UASs), or other gene features. It is also unclear whether UASs can broadly activate transcription from the different promoter classes or whether efficient transcription requires matching UASs and promoters of similar gene class. Here we measure transcription and cofactor specificity for tens of thousands of UAS-core promoter combinations. We find that <5% of UASs display strong core promoter specificity while most UASs can broadly activate promoters regardless of regulatory class. However, we find that matching UASs and promoters from the same gene class is generally important for optimal expression. From examining the cofactor dependence of this large UAS-promoter set, we find that sensitivity to rapid depletion of MED Tail or SAGA is dependent on the identity of both UAS and promoter while dependence on TFIID localizes to only the core promoter. Our results explain why transcription factor-mediated MED recruitment to the UAS does not always result in Tail-dependent transcription and highlight the role of TATA and TATA-like promoter sequences in MED Tail function.

molecular biology↗

bioRxiv2022

Proximal dendritic localization of NALCN channels underlies tonic and burst firing in nigral dopaminergic neurons

In multipolar nigral dopamine (DA) neurons, the highly excitable proximal dendritic compartments (PDCs) and two Na+-permeable leak channels, TRPC3 and NALCN, play a key role in pacemaking. However, the causal link between them is unknown. Here we report that the proximal dendritic localization of NALCN underlies pacemaking and burst firing in DA neurons. Our morphological analysis of nigral DA neurons reveals that TRPC3 is ubiquitously expressed in the whole somatodendritic compartment, but NALCN is localized within the PDCs. Blocking either TRPC3 or NALCN channels abolished pacemaking. However, only blocking NALCN, not TRPC3, degraded burst discharges. Furthermore, local glutamate uncaging readily induced burst discharges within the PDCs, compared with other parts of the neuron, and NALCN channel inhibition dissipated burst generation, indicating the importance of NALCN to the high excitability of PDCs. Therefore, we conclude that PDCs serve as a common base for tonic and burst firing in nigral DA neurons.

neuroscience↗

bioRxiv2022

Longitudinal assessment of brain structure and behavior in youth with rapid weight gain: Potential contributing causes and consequences

ObjectiveIndependent of weight status, rapid weight gain has been associated with underlying brain structure variation in regions associated with food intake and impulsivity among pre-adolescents. Yet, we lack clarity on how developmental maturation coincides with rapid weight gain and weight stability. MethodsWe identified brain predictors of two-year rapid weight gain and its longitudinal effects on brain structure and impulsivity in the Adolescent Brain Cognitive DevelopmentSM Study(R). Youth were categorized as Healthy Weight/Weight Stable (WSHW, n=225) or Weight Gainers (WG, n=221, >38lbs); 63% of the WG group were healthy weight at 9-to-10-years-old. ResultsA five-fold cross-validated logistic elastic-net regression revealed that rapid weight gain was associated with structural variation amongst 39 brain features at 9-to-10-years-old in regions involved with executive functioning, appetitive control, and reward sensitivity. Two years later, WG youth showed differences in change over time in several of these regions and performed worse on measures of impulsivity. ConclusionsThese findings suggest that brain structure in pre-adolescence may predispose some to rapid weight gain and that weight gain itself may alter maturational brain change in regions important for food intake and impulsivity. Behavioral interventions that target inhibitory control may improve trajectories of brain maturation and facilitate healthier behaviors.

neuroscience↗

bioRxiv2021

Mediator is broadly recruited to gene promoters via a Tail-independent mechanism

Mediator (MED) is a conserved factor with important roles in both basal and activated transcription. It is believed that MED plays a direct role in transcriptional regulation at most genes by functionally bridging enhancers and promoters. Here, we investigate the genome-wide roles of yeast MED by rapid depletion of its activator-binding domain (Tail) and monitoring changes in nascent transcription. We find that MED Tail and activator-mediated MED recruitment regulate only a small subset of genes. At most genes, MED bypasses the UAS and is directly recruited to promoters to facilitate transcription initiation. Our results define three classes of genes that differ in PIC assembly pathways and the requirements for MED Tail, SAGA, TFIID and BET factors Bdf1/2. We also find that the depletion of the MED middle module subunit Med7 mimics inactivation of Tail, suggesting a functional link. Our combined results have broad implications for the roles of MED, other coactivators, and mechanisms of transcriptional regulation at different gene classes.

molecular biology↗

bioRxiv2021

Performance Scaling for Structural MRI Surface Parcellations: A Machine Learning Analysis in the ABCD Study

The use of pre-defined parcellations on surface-based representations of the brain as a method for data reduction is common across neuroimaging studies. In particular, prediction-based studies typically employ parcellation-driven summaries of brain measures as input to predictive algorithms, but the choice of parcellation and its influence on performance is often ignored. Here we employed pre-processed structural magnetic resonance imaging data (sMRI) from the ABCD Study(R) to examine the relationship between 220 parcellations and out-of-sample predictive performance across 45 phenotypic measures in a large sample of 9-10-year-old children (N=9,432). Choice of Machine Learning (ML) pipeline and use of alternative multiple parcellation-based strategies were also assessed. Relative parcellation performance was dependent on the spatial resolution of the parcellation, with larger numbers of parcels (up to [~]4000) outperforming coarser parcellations, according to a power-law scaling of between 1/4 and 1/3. Performance was further influenced by the type of parcellation, ML pipeline, and general strategy, with existing literature-based parcellations, a support vector based pipeline, and ensembling across multiple parcellations, respectively, as the highest performing. These findings highlight the choice of parcellation as an important influence on downstream predictive performance, showing in some cases that switching to a higher resolution parcellation can yield a relatively large boost to performance.

neuroscience↗

bioRxiv2021

Circulatory neutrophils exhibit enhanced neutrophil extracellular trap formation in early puerperium: NETs at the nexus of thrombosis and immunity?

Pregnancy is associated with elevated maternal levels of cell-free DNA of neutrophil extracellular trap (NET) origin, as circulatory neutrophils exhibit increased spontaneous NET formation, mainly driven by G-CSF and finely modulated by sex hormones. The postpartum period, on the other hand, involves physiological alterations consistent with the need for protection against infections and fatal haemorrhage. Our findings indicate that all relevant serum markers of neutrophil degranulation and NET release are substantially augmented postpartum. Neutrophil pro-NETotic activity in vitro is also upregulated particularly in post-delivery neutrophils. Moreover, maternal puerperal neutrophils exhibit a strong pro-NETotic phenotype, associated with increased levels of all key players in the generation of NETs - citH3, MPO, NE and ROS, compared to non-pregnant and pregnant controls. Intriguingly, post-delivery NET formation is independent of G-CSF in contrast to late gestation and complemented by the presence of TF on the NETs, alterations in the platelet activity status, and activation of the coagulation cascade, triggered by circulating microparticles. Taken together, our results reveal the highly pro-NETotic and potentially procoagulant nature of postpartum neutrophils, bridging an overt immune activation with possible harmful thrombotic incidence.

immunology↗

bioRxiv2021

Cooperation between CYB5R3 and NOX4 via coenzyme Q mitigates endothelial inflammation

NADPH oxidase 4 (NOX4) regulates endothelial inflammation by producing reactive oxygen species. Since coenzyme Q (CoQ) mimics affect NOX4 activity, we hypothesize that cytochrome b5 reductase 3 (CYB5R3), a CoQ reductase abundant in vascular endothelial cells, modulates inflammatory activation. Mice lacking endothelial CYB5R3 (R3 KO), under lipopolysaccharides (LPS) challenge, showed exacerbated hypotension, decreased acetylcholine-induced vasodilation, and elevated vascular adhesion molecule 1 (Vcam-1) mRNA in aorta. In vitro, silencing Cyb5r3 enhanced LPS-induced VCAM-1 protein in a NOX4 dependent manner. APEX2- based electron microscopy and proximity biotinylation demonstrated CYB5R3s localization on the mitochondrial outer membrane and its interaction with NOX4, which was further confirmed by the proximity ligation assay. Notably, Cyb5r3 silenced HAECs had less total H2O2 but more mitochondrial O2*-. Using inactive or non-membrane bound active CYB5R3, we found CYB5R3 activity and membrane translocation were needed for optimal generation of H2O2 by NOX4. Lastly, CoQ deficient cells showed decreased NOX4-derived H2O2, indicating a requirement for endogenous CoQ in NOX4 activity. In conclusion, CYB5R3 mitigates endothelial inflammatory activation by assisting in NOX4-dependent H2O2 generation via CoQ. NOVELTY AND SIGNIFICANCEO_ST_ABSWhat Is Known?C_ST_ABSNADPH oxidase 4 (NOX4) reportedly produces primarily hydrogen peroxide (H2O2) and, to a lesser extent, superoxide (O2*-) and has been shown to have both beneficial and deleterious effects in the cardiovascular system. NOX4 activity can be affected by NAD(P)H quinone oxidoreductase 1 (NQO1), a CoQ reductase, and synthetic quinone compounds used to mimic CoQ. Cytochrome b5 reductase 3 (CYB5R3) is known to reduce CoQ and is highly expressed in endothelial cells. What New Information Does This Article Contribute?In vivo, the lack of endothelial CYB5R3 causes exacerbated lipopolysaccharides (LPS)-induced inflammatory signaling, endothelial dysfunction, and hypotension. Endothelial CYB5R3 mitigates inflammatory signaling by LPS and tumor necrosis factor (TNF-) in a NOX4 dependent manner. In endothelial cells, CYB5R3 and NOX4 reside in close proximity on the mitochondrial outer membrane. NOX4s ability to generate H2O2 depends on the membrane translocation and activity of CYB5R3 and the presence of endogenous CoQ. NONSTANDARD Abbreviations and Acronyms [Table 1] Protein names are abbreviated as capital letters (e.g., CYB5R3), while the corresponding gene names are annotated as in italic lower cases (e.g., Cyb5r3).

cell biology↗

bioRxiv2021

Functional metabolic phenotyping of human pancreatic ductal adenocarcinoma

Pancreatic Ductal Adenocarcinoma (PDAC) lacks targeted treatment options. Although subtypes with transcriptome-based distinct lineage and differentiation features have been identified, deduced clinically actionable targets remain elusive. We here investigate functional metabolic features of the classical and QM (quasi-mesenchymal)/basal-like PDAC subtypes potentially exploitable for non-invasive subtype differentiation and therapeutic intervention. A collection of human PDAC cell lines, primary patient derived cells (PDC), patient derived xenografts (PDX) and patient PDAC samples were transcriptionally stratified into the classical and QM subtype. Functional metabolic analyses including targeted and non-targeted metabolite profiling (matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI)), seahorse metabolic flux assays and metabolic drug targeting were performed. Hyperpolarized 13C-magnetic resonance spectroscopy (HP-MRS) of PDAC xenografts was used for in vivo detection of intra-tumoral [1-13C]pyruvate and [1-13C]lactate metabolism. We identified glycolysis and lipid metabolism/fatty acid oxidation as transcriptionally preserved metabolic pathways in QM and classical PDAC subtype respectively. However, these metabolic cues were not unambiguously functionally linked to one subtype. Striking functional metabolic heterogeneity was observed especially in primary patient derived cells with only individual samples representing high dependence on glycolysis or mitochondrial oxidation. Of note, QM cells actively use the glycolytic product lactate as oxidative mitochondrial fuel. Using HP-MRS, we were able to non-invasively differentiate glycolytic tumor xenografts with high intratumoral [1-13C]pyruvate to [1-13C]lactate conversion in vivo. Although PDAC transcriptomes indicate molecular subtype-associated distinct metabolic pathways, we found substantial functional metabolic heterogeneity independent of the molecular subtype. Non-invasive identification of highly glycolytic tumors by [1-13C]pyruvate/lactate HP-MRS support individualized metabolic targeting approaches.

cancer biology↗

bioRxiv2021

Suv420 enrichment at the centromere limits Aurora B localization and function

Centromere structure and function are defined by the epigenetic modification of histones at centromeric and pericentromeric chromatin. The constitutive heterochromatin found at pericentromeric regions is highly enriched for H3K9me3 and H4K20me3. While mis-expression of the methyltransferase enzymes, Suv39 and Suv420, that regulate these marks are common in disease, the consequences of such changes are not well understood. Our data show that increased centromere localization of Suv39 and Suv420 suppress centromere transcription and compromise localization of the mitotic kinase Aurora B: decreasing microtubule dynamics and compromising chromosome alignment and segregation. We find that inhibition of Suv420 methyltransferase activity partially restores Aurora B localization to centromeres and that restoration of the Aurora B-containing CPC to the centromere is sufficient to suppress mitotic errors that result when Suv420/H4K20me3 is enriched at centromeres. Consistent with a role for Suv39 and Suv420 in negatively regulating Aurora B, high expression of these enzymes corresponds with increased sensitivity to Aurora kinase inhibition in cancer cells suggesting that increased H3K9 and H4K20 methylation may be an underappreciated source of chromosome missegregation in cancer.

cell biology↗

bioRxiv2021

TRPC3 and NALCN channels drive pacemaking in substantia nigra dopaminergic neurons

Midbrain dopamine (DA) neurons are slow pacemakers that maintain extracellular DA levels. During the interspike intervals, subthreshold slow depolarization underlies autonomous pacemaking and determines its rate. However, the ion channels that determine slow depolarization are unknown. Here we show that TRPC3 and NALCN channels together form sustained inward currents responsible for the slow depolarization of nigral DA neurons. Specific TRPC3 channel blockade completely blocked DA neuron pacemaking, but the pacemaking activity in TRPC3 knock-out (KO) mice was perfectly normal, suggesting the presence of compensating ion channels. Blocking NALCN channels abolished pacemaking in both TRPC3 KO and wild-type mice. The NALCN current and mRNA and protein expression are increased in TRPC3 KO mice, indicating that NALCN compensates for TRPC3 currents. In normal conditions, TRPC3 and NALCN contribute equally to slow depolarization. Therefore, we conclude that TRPC3 and NALCN are two major leak channels that drive robust pacemaking in nigral DA neurons.

neuroscience↗

bioRxiv2021

BET family members Bdf1/2 modulate global transcription initiation and elongation in Saccharomyces cerevisiae

Human bromodomain-containing BET family members are promising targets for therapy of cancer and immunoinflammatory diseases, but their mechanisms of action and functional redundancies are poorly understood. Bdf1/2, yeast homologues of the human BET factors, were previously proposed to target transcription factor TFIID to acetylated histone H4, analogous to bromodomains that are present within the largest subunit of metazoan TFIID. We investigated the genome-wide roles of Bdf1/2 and found that their important contributions to transcription extend beyond TFIID function, as transcription of many genes is more sensitive to Bdf1/2 than to TFIID depletion. Bdf1/2 co-occupy the majority of yeast promoters and affect preinitiation complex formation through recruitment of TFIID, Mediator and basal transcription factors to chromatin. Surprisingly, we discovered that hypersensitivity of genes to Bdf1/2 depletion results from combined defects in transcription initiation and processive elongation, a striking functional similarity to human BET proteins, most notably Brd4. Our results establish Bdf1/2 as critical for yeast transcription and provide important mechanistic insights into the function of BET proteins in all eukaryotes.

molecular biology↗

bioRxiv2021

An improved ChEC-seq method accurately maps the genome-wide binding of transcription coactivators and sequence-specific transcription factors

Mittal and colleagues have raised questions about mapping transcription factor locations on DNA using the MNase-based ChEC-seq method (Mittal et al., 2021). Partly due to this concern, we modified the experimental conditions of the MNase cleavage step and subsequent computational analyses, resulting in more stringent conditions for mapping protein-DNA interactions (Donczew et al., 2020). The revised method (dx.doi.org/10.17504/protocols.io.bizgkf3w) answers questions raised by Mittal et al. and, without changing earlier conclusions, identified widespread promoter binding of the transcription coactivators TFIID and SAGA at active genes. The revised method is also suitable for accurately mapping the genome-wide locations of DNA sequence-specific transcription factors.

genomics↗

bioRxiv2020

The role of XPB/Ssl2 double-stranded DNA translocase processivity in transcription start-site scanning.

The general transcription factor TFIIH contains three ATP-dependent catalytic activities. TFIIH functions in nucleotide excision repair primarily as a DNA helicase and in Pol II transcription initiation as a dsDNA translocase and protein kinase. During initiation, the XPB/Ssl2 subunit of TFIIH couples ATP hydrolysis to dsDNA translocation facilitating promoter opening and the kinase module phosphorylates the C-terminal domain to facilitate the transition to elongation. These functions are conserved between metazoans and yeast; however, yeast TFIIH also drives transcription start-site scanning in which Pol II scans downstream DNA to locate productive start-sites. The ten-subunit holo-TFIIH from S. cerevisiae has a processive dsDNA translocase activity required for scanning and a structural role in scanning has been ascribed to the three-subunit TFIIH kinase module. Here, we assess the dsDNA translocase activity of ten-subunit holo- and core-TFIIH complexes (i.e. seven subunits, lacking the kinase module) from both S. cerevisiae and H. sapiens. We find that neither holo nor core human TFIIH exhibit processive translocation, consistent with the lack of start-site scanning in humans. Furthermore, in contrast to holo-TFIIH, the S. cerevisiae core-TFIIH also lacks processive translocation and its dsDNA-stimulated ATPase activity was reduced ~5-fold to a level comparable to the human complexes, potentially explaining the reported upstream shift in start-site observed in the absence of the S. cerevisiae kinase module. These results suggest that neither human nor S. cerevisiae core-TFIIH can translocate efficiently, and that the S. cerevisiae kinase module functions as a processivity factor to allow for robust transcription start-site scanning.

biochemistry↗