Biology subjects

Steinberg, M.

Publications and source records attributed to Steinberg, M..

4 recordsLinked to original sources

bioRxiv2026

ACHT2 deactivates carbon assimilation during light-dark transition

In chloroplasts, photosynthetic efficiency relies on a delicate balance between reductive and oxidative thiol-based signaling networks. Members of the high-midpoint-potential atypical thioredoxins (Trxs) were shown to oxidize photosynthetic enzymes by channeling reducing equivalents to H2O2 through 2-Cys-Prx activity. However, it remains unclear which atypical-Trx isoforms regulate Calvin-Benson cycle (CBC) inactivation, and whether they possess distinct functional specificities or operate redundantly in vivo. To resolve this, electron transport and carbon assimilation were continuously monitored during physiological dynamic light transitions in CRISPR-generated single, double and triple atypical Trx mutants. Notably, ACHT2 was identified as a primary determinant of CBC inactivation during dark-to-light transitions, as evidenced by the alleviation of redox-mediated bottlenecks in electron flow downstream of Fd and a lower CBC inactivation state in acht2 plants, resembling the phenotype observed in plants lacking 2-Cys Prxs A and B (2cpab). In contrast, plants mutated in ACHT1, ACHT4, or TrxL2 displayed CBC inactivation kinetics comparable to those of the wild type. Furthermore, mutation of ACHT2 did not compromise plant fitness. In contrast, growth retardation was observed in the acht1/acht4 double mutant, suggesting that the severe phenotype of 2cpab does not arise from impaired CBC inactivation, but rather from disruption of oxidative regulation of other metabolic pathways mediated by distinct atypical Trxs. Collectively, these findings reveal a high degree of regulatory specificity within the chloroplast oxidative network and provide a foundation for a deeper understanding of how activation-inactivation cycles contribute to plant adaptation to dynamic light environments.

plant biology↗

bioRxiv2026

Dark-induced inactivation of the carbon assimilation process requires a water water cycle driven oxidative burst

Chloroplast metabolism is constantly fine-tuned by light availability through the perception and transmission of reductive and oxidative signals that activate or deactivate distinct metabolic enzymes. The reducing power originating from the photosynthetic electron transport chain has been shown to fuel the redox regulatory network, linking electron transport to the reductive activation of photosynthetic enzymes. However, the source of the oxidizing equivalents required to reverse photosynthetic enzyme activation and drive them toward an oxidized, inactive state has not yet been experimentally demonstrated. Here, we resolve redox dynamics associated with carbon assimilation inactivation by combining time-resolved redox imaging during the light-to-dark transition (LDT) with gas-exchange-based measurements. Dark-induced inactivation of carbon assimilation proved oxygen-dependent and coincided with an oxygen-dependent oxidative burst triggered during the LDT. This oxidative burst was suppressed under conditions that blocked electron transport to PSI or in plants in which PSI was photoinactivated. Notably, pgr5 and pgrl1ab mutants exhibited attenuated oxidative bursts and suppressed LDT-associated carbon assimilation inactivation, demonstrating that PGR5/PGRL1-dependent activity is required to generate the oxidative burst that drives CBC inactivation during LDT. These results establish a direct mechanistic link between oxygen- and PSI-dependent oxidative bursts and the inhibition of photosynthesis and mark the water-water cycle (WWC) as the primary source of the transient accumulation of oxidative equivalents that drive inactivation of Calvin-Benson cycle enzymes in darkness.

plant biology↗

bioRxiv2025

Comprehensive cellular analysis with single-nucleus RNA-seq of archived PAXgene whole blood samples

No method has been developed for single-cell analysis of the large repositories of preserved whole blood samples stored in PAXgene Blood RNA tubes. To address this gap, two nuclei isolation techniques for single-nucleus RNA sequencing were evaluated: mechanical separation (MS), using an Acrodisc filter, and cell lysis (CL). While both methods captured nuclei from all major immune cell types, CL resulted in up to two orders of magnitude higher nuclei yields and less biased proportions of immune cells than MS. High ambient globin gene counts following CL were substantially reduced by CRISPR-guided globin gene depletion of complementary DNA, resulting in more sensitive and efficient gene detection per cell. Despite capturing only nuclear transcripts, CL-derived samples maintained similar cell-type proportions and gene expression as matched peripheral blood mononuclear cell samples, while retaining granulocytes. The CL isolation with globin depletion method enables comprehensive analysis of PAXgene whole blood samples at single-cell resolution. MOTIVATIONPAXgene Blood RNA Tubes are commonly used for blood preservation and gene expression studies due to the ease of use and stabilization of RNA. To date, transcriptomic studies using PAXgene tubes have been limited to bulk RNA profiling approaches such as RNA sequencing (RNA-seq). Here we introduce a method for recovering high-quality nuclei from frozen blood preserved in PAXgene tubes for single-nucleus RNA-seq (snRNA-seq). Using this approach, all white blood cell types - including granulocytes - are captured, thereby enabling comprehensive transcriptomic profiling of peripheral blood at single-cell resolution.

genomics↗

bioRxiv2020

Human papillomavirus 16 positive cervical cancer in Guatemala: The D2 and D3 sublineages differ in integration rate and age of diagnosis

Human papillomavirus (HPV) 16 displays substantial sequence variation; four HPV16 lineages (A, B, C, D) have been described, as well as multiple sub-lineages. To identify molecular events associated with HPV16 carcinogenesis we evaluated viral variation, the integration of HPV16, and somatic mutation in 96 cervical cancer samples from Guatemala. A total of 64% (60/94) of the samples had integrated HPV16 sequences, and integration was associated with an earlier age of diagnosis (P=0.0007) and pre-menopausal disease. HPV16 integration sites were broadly distributed in the genome but in one tumor, HPV16 integrated into the promoter of the interferon regulatory factor 4 (IRF4) gene, which plays an important role in the regulation of the interferon response to viral infection. The HPV16 D2 and D3 sub-lineages were found in 23% and 30% of the tumors, respectively and were significantly associated with adenocarcinoma. D2-positive tumors had a higher rate of integration (P=0.011), earlier age of diagnosis (P=0.012), and a lower rate of somatic mutation (P=0.03). Whereas D3-positive tumors are less likely to integrate, have later age-of-diagnosis, and a higher rate of somatic mutation. In conclusion, Guatemalan cervical tumors have a high frequency of the very high-risk HPV16 D2 and D3 sub-lineages and cervical cancer patients with these variants of HPV16 differ in histology, age of- diagnosis, integration, and somatic mutation frequency. In summary, related lineages of HPV16 have different features of oncogenicity.

cancer biology↗