Biology subjects

Brandt, K.

Publications and source records attributed to Brandt, K..

6 recordsLinked to original sources

bioRxiv2026

A unified photosensitizer platform for in situ DNA, RNA, and protein directed proximity labeling

Cells depend on the spatial organization of proteins, RNA, and DNA into discrete subcellular compartments. Previous methods have largely centered on measuring spatial organization based on only one of these biomolecular classes at a time. Here, we demonstrate that POCA photocatalytic proximity labeling can serve as a unified photosensitizer-based platform for profiling the proximal proteomes of protein, RNA, and DNA targets within a single experimental framework. We show that POCA can harness standard immunofluorescence or in situ hybridization workflows to specifically target organic fluorophore photosensitizers to intracellular targets for proximity labeling in fixed cells. POCA-targeted proximity labeling requires minimal cellular input and does not require genetic engineering. Additionally, POCA photosensitizers are selected to also be fluorescent, enabling direct confirmation of on-target localization by imaging prior to proteomic analysis. To demonstrate broad utility, we apply POCA across multiple molecular targets spanning protein, RNA, and genomic DNA, including components of the nuclear pore complex, nucleolus, nuclear speckles, telomeres, and pericentromeric heterochromatin. By anchoring proximity labeling to both a protein and an RNA within the same nuclear compartment, we resolve shared and distinct proximal proteomes from orthogonal molecular perspectives.

cell biology↗

bioRxiv2026

Tissue composition shapes differential skeletal integration strategies during axolotl limb regeneration

Limb regeneration requires rebuilding the missing structures and integrating them with stump tissues. Osteoclast-mediated tissue resorption is essential for skeletal integration during regeneration. However, it is unknown if skeletal tissue composition impacts its resorption and, if so, how it is regulated. Here, we show that osteoclast-mediated skeletal resorption is activated in amputations damaging the diaphysis, but not the epiphysis, of skeletal elements. We found that amputations in the calcified diaphysis trigger the sustained expression of RANKL and the chemokine Loc483, and that the latter is sufficient to induce osteoclast presence in non-resorbing amputations. Finally, our data suggests that the transcriptomic profile of the apical ectodermal cap is impacted by the tissues injured by the amputation. Overall, our work reveals that tissue composition at the amputation plane shapes adaptations of the regenerative program, which ultimately contribute to the near-seamless tissue integration of the regenerating axolotl limb regardless of the amputation position.

developmental biology↗

bioRxiv2024

Vasoactive Intestinal Peptide Amphiphile Micelle Material Properties Influence Their Cell Association and Internalization

Vasoactive intestinal peptide (VIP) is a promising anti-inflammatory peptide therapeutic that is known to induce biological effects by interacting with its cognate receptor (i.e., VPAC) on the surface of antigen presenting cells (APCs). For VIP-based drug delivery technologies like nano- and microparticles, little is known about the effect VPAC targeting has on APC behavior. This is further influenced by the fact that particulate material properties including chemistry, shape, and size are all known to influence APC behavior. In this study, peptide amphiphile micelles (PAMs) were employed as a modifiable platform to study the impact VPAC targeting and physical particle properties have on their association with macrophages. VIP amphiphile micelles (VIPAMs) and their scrambled peptide amphiphile micelle analogs (SVIPAMs) were fabricated from various chemistries yielding particle batches that were comprised of spheres (10 - 20 nm in diameter) and/or cylinders of varying lengths (i.e., 20 - 9000 nm). Micelle surface attachment to and internalization by macrophages were observed using confocal microscopy and their association was characterized by flow cytometry. The enclosed work provides strong evidence that macrophages rapidly bind VPAC specific micelles independent of physical properties though micelle shape and size as well as receptor-specificity all influence their long-term macrophage association. Specifically, a mixture of spherical and short cylindrical VIPAMs were able to achieve the greatest cell association which may correlate to their capacity to fully bind the VPAC receptors available on the surface of macrophages. These results provide the foundation of how nano- and microparticle physical properties and targeting capacity synergistically influence their capacity to associate with APCs.

bioengineering↗

bioRxiv2023

Technical and biological sources of unreliability of Infinium type II probes of the Illumina MethylationEPIC BeadChip microarray

The Illumina Methylation array platform has facilitated countless epigenetic studies on DNA methylation (DNAme) in health and disease, yet relatively few studies have so studied its reliability, i.e., the consistency of repeated measures. Here we focus on the reliability of both type I and type II Infinium probes. We propose a method for excluding unreliable probes based on dynamic thresholds for mean intensity (MI) and unreliability, estimated by probe-level simulation of the influence of technical noise on methylation {beta}-values using the background intensities of negative control probes. We validate our method in several datasets, including Illumina MethylationEPIC BeadChip v1.0 data from paired whole blood samples taken six weeks apart. Our analysis revealed that specifically probes with low MI exhibit higher {beta}-value variability between repeated samples. MI was associated with the number of C-bases in the respective probe sequence and correlated negatively with unreliability scores. The unreliability scores were substantiated through validation in a new EPIC v1.0 (blood and cervix) and a publicly available 450k (blood) dataset, as they effectively captured the variability observed in {beta}-values between technical replicates. Finally, despite promising higher robustness, the newer version v2.0 of the MethylationEPIC BeadChip retained a substantial number of probes with poor unreliability scores. To enhance current pre-processing pipelines, we developed an R package to calculate MI and unreliability scores and provide guidance on establishing optimal dynamic score thresholds for a given data set.

genomics↗

bioRxiv2023

Multi-species atlas resolves an axolotl limb development and regeneration paradox

Humans and other tetrapods are considered to require apical-ectodermal-ridge, AER, cells for limb development, and AER-like cells are suggested to be re-formed to initiate limb regeneration. Paradoxically, the presence of AER in the axolotl, the primary regeneration model organism, remains controversial. Here, by leveraging a single-cell transcriptomics-based multi-species atlas, composed of axolotl, human, mouse, chicken, and frog cells, we first established that axolotls contain cells with AER characteristics. Surprisingly, further analyses and spatial transcriptomics revealed that axolotl limbs do not fully re-form AER cells during regeneration. Moreover, the axolotl mesoderm displays part of the AER machinery, revealing a novel program for limb (re)growth. These results clarify the debate about the axolotl AER and the extent to which the limb developmental program is recapitulated during regeneration.

developmental biology↗

bioRxiv2019

Neuron-glia interaction through Serotonin-BDNF-NGFR axis enables regenerative neurogenesis in Alzheimer’s model of adult zebrafish brain

It was recently suggested that supplying the brain with new neurons could counteract Alzheimers disease. This provocative idea requires further testing in experimental models where the molecular basis of disease-induced neuronal regeneration could be investigated. We previously found that zebrafish stimulates neural stem cell (NSC) plasticity and neurogenesis in Alzheimers disease and could help to understand the mechanisms to be harnessed for develop new neurons in diseased mammalian brains. Here, by performing single-cell transcriptomics, we found that Amyloid toxicity-induced Interleukin-4 induces NSC proliferation and neurogenesis by suppressing the tryptophan metabolism and reducing the production of Serotonin. NSC proliferation was suppressed by Serotonin via downregulation of BDNF-expression in Serotonin-responsive periventricular neurons. BDNF enhances NSC plasticity and neurogenesis via NGFRA/NFkB signaling in zebrafish but not in rodents. Collectively, our results suggest a complex neuron-glia interaction that regulates regenerative neurogenesis after Alzheimers disease conditions in zebrafish.\n\nKey findings- Amyloid-induced Interleukin-4 suppresses Serotonin (5-HT) production in adult zebrafish brain\n- 5-HT affects htr1-expresing neurons and suppresses bdnf expression\n- BDNF enhances plasticity in neural stem cells via NGFRA/NFkB signaling\n- BDNF/NGFRA signaling is a neuro-regenerative mechanism in zebrafish but not in mammals.

neuroscience↗