Journal: bioRxiv

Article Title: Single-Cell Profiling of the Developing Organ of Corti Identifies Etv4/5/1 as Key Regulators of Pillar Cell Identity

doi: 10.64898/2026.01.19.700450

Figure Lengend Snippet: a. At E18, PROX1 + -IPCs, OPCs and DCs are arranged in ordered rows in the basal region of the control cochlea. In the less mature apical region (lower panel) the lateral rows of DCs are not completely aligned yet. In contrast, in an Etv Triple KO cochlea overall cellular alignment is disrupted in both the basal and apical regions. a’. In a control cochlea, OPCs (CD44 + , magenta) form a single ordered row in the basal region but are only weakly CD44 + in the apex. In an Etv Triple KO cochlea multiple rows of CD44 + -OPCs are present in the basal and apical regions. b. Cross-sectional Z-stacks of the OC from the basal region of a control and an Etv Triple KO. In the control, the single CD44 + /PROX1 + -OPC (arrow) is located adjacent to the first OHC (numbered). In the Etv Triple KO, three OPCs (arrows), are intermixed with the first two rows of OHCs (numbered). c. The positions of the six nearest PROX1 + -nuclei were mapped relative to fourth row PROX1 + -cells (see text for details). The radar plot illustrates results for control and Etv Triple KOs. In control OCs, the spatial alignment of PROX1 + -DCs is regular (green profile). In contrast, in Etv Triple KOs spatial patterning is disrupted (magenta). d . Surface view of the OC in control and Etv Triple KOs at E18 labeled with anti-S100A1, a marker of IHCs, PCs, and DCs, and phalloidin. In controls, DCs (arrows) form dumbbell shapes between OHCs. Although there are some patterning defects in the Etv Triple KO OC (arrows), overall expression of S100A1 appears comparable to control. S100A1-labeling of IHCs was also present in Etv Triple KOs (not shown). e. Surface views of an E18 control OC illustrating all HCs (magenta) and OHCs (POU4F3 + (magenta) and BCL11B + (green)) and a similar view from an Etv Triple KO showing an over-production of HCs in the lateral region of the OC(numbered), many of which are negative for BCL11B (arrows). f-f’’. Cross-sectional Z-stack views of the OC from a control and an Etv Triple KO at E18. In the control, a single IHC (arrow), three OHCs (numbered), a single IPC and OPC, and three DCs are present. In an Etv Triple KO, an ectopic PROX1 + -HC (asterisk) is present in the pillar cell region. g-g’’. Cross-sectional Z-stack views as in g but labeled with MYO6 instead of POU4F3. In the Etv Triple KO, a PROX1 + /MYO6 + -cell is present in the PC region of the OC (arrowhead and asterisk). Inset: Image of the ectopic hair cell (asterisk) in gray scale with increased contrast for improved detection of MYO6 labeling. h. Surface views of the basal region of the OC from animals with the indicated genotypes at E16. In controls a single row of NGFR + -IPCs (magenta) is already present. Some IPCs are also positive for NPY (green). In contrast, in an Etv4/5/1 Triple KO only minimal expression of NGFR is present while NPY expression is absent. In addition, ectopic HCs (arrows) are present in the pillar cell space. i. Surface view of the basal region of the OC from control and an Etv Triple KO at E16. In the control a single row of MYO6 + -IHCs and three rows of MYO6 + /BCL11B + -OHCs are present. In an Etv Triple KO only three rows of OHCs are present by comparison with the four rows present in some regions at E18 . Some PROX1 + -nuclei are present in the HC nuclear layer (arrows) but are negative for HC markers. Scale bar in a,b,e,f,g (same in h), and j (same in i), 20 μm.

Article Snippet: The primary antibodies used were rabbit anti-MYOSIN6 (Proteus Biosciences 25-6791, RRID:AB_10013626 at 1:500 dilution), rabbit anti-NPY (Peninsula Laboratories T-4070, RRID: AB_518504 at 1:500 dilution), goat anti-NGFR (R&D Systems AF367, RRID:AB_2152638 at 1:500 dilution), goat anti-PROX1 (R&D Systems AF2727, RRID:AB_2170716 at 1:500 dilution), goat anti-SOX2 (R&D Systems AF2018, RRID:AB_355110 at 1:500 dilution), rat anti-CD44 (BD biosciences 550538, RRID: AB_393732 at 1:500 dilution), mouse anti-POU4F3 (Santa Cruz biotechnology, RRID: AB_2167543 at 1:200 dilution), goat anti-MYOSIN6 (Kelley lab custom made at 1:500 dilution), mouse anti-CTIP2 (BCL11B) (Abcam ab18465, RRID: AB_2064130 at 1:500 dilution), rabbit anti-NRP2 (Cell Signaling D39A5, AB_2155250 at 1:500), sheep anti-S100A1 (R&D Systems, AF4476, AB_2183326 at 1:500).

Techniques: Control, Labeling, Marker, Expressing, Comparison

Journal: ACS Chemical Neuroscience

Article Title: Calcium-Dependent S100A8 Amyloid Fibril Formation via S100A1-Mediated Transient Interaction

doi: 10.1021/acschemneuro.5c00086

Figure Lengend Snippet: (A) During neuroinflammation (1), astrocytes upregulate the expression of S100A1, S100A8, and other S100 proteins. These proteins are secreted via noncanonical pathways (2) and transfer inflammation signals to glial or neuronal cells (3). S100 proteins can accumulate and contribute to the formation of corpora amylacea (4) and amyloid plaques (5). (B) Transcriptional differences of S100 RNA in various brain regions according to The Human Protein Atlas (nTPM, transcripts per million).

Article Snippet: S100A1 protein is highly prevalent in the heart; however, it is also located in the brain, rivaling transcriptional levels to those of S100B, according to The Human Protein Atlas ( B), whereas S100A8 and S100A9 expression rises during neuroinflammation.

Techniques: Expressing

Journal: ACS Chemical Neuroscience

Article Title: Calcium-Dependent S100A8 Amyloid Fibril Formation via S100A1-Mediated Transient Interaction

doi: 10.1021/acschemneuro.5c00086

Figure Lengend Snippet: Aggregation and stability propensities of S100A1 and S100A8 proteins. (A) Aggregation kinetics of S100A1, S100A8, and S100A1/S100A8 in the presence or absence of calcium ions, followed by ThT fluorescence. (B) Maximum ThT fluorescence values of aggregation kinetics. (C) Densitometric quantification of filtered S100A1, S100A8, and S100A1/S100A8 samples after aggregation. (D) Chaperone assay of S100A1 (10 μM), S100A8 (10 μM), and S100A1/S100A8 (10 μM each) against DTT-induced aggregation of lysozyme (0.2 mg/mL). (E) Melting temperatures of S100A1, S100A8, and S100A1/S100A8 proteins.

Article Snippet: S100A1 protein is highly prevalent in the heart; however, it is also located in the brain, rivaling transcriptional levels to those of S100B, according to The Human Protein Atlas ( B), whereas S100A8 and S100A9 expression rises during neuroinflammation.

Techniques: Fluorescence

Journal: ACS Chemical Neuroscience

Article Title: Calcium-Dependent S100A8 Amyloid Fibril Formation via S100A1-Mediated Transient Interaction

doi: 10.1021/acschemneuro.5c00086

Figure Lengend Snippet: Morphology and colocalization of S100A1 and S100A8 aggregates. AFM images of (A) S100A1, S100A8, and (B) S100A1/S100A8 after 70 h of aggregation (scale bar: 500 nm). (C) The height distribution of S100A1/S100A8 samples with box plots indicating the median, interquartile range (IQR), and whiskers 1.5× range of IQR from box. (D) Transmission electron microscopy (scale bar: 500 nm) and (E) fluorescence microscopy (scale bar: 10 μm) images of S100A1/S100A8 aggregated mixtures at different calcium ion concentrations. Additional fluorescence images are presented in Figures S3–S5 .

Article Snippet: S100A1 protein is highly prevalent in the heart; however, it is also located in the brain, rivaling transcriptional levels to those of S100B, according to The Human Protein Atlas ( B), whereas S100A8 and S100A9 expression rises during neuroinflammation.

Techniques: Transmission Assay, Electron Microscopy, Fluorescence, Microscopy

Journal: ACS Chemical Neuroscience

Article Title: Calcium-Dependent S100A8 Amyloid Fibril Formation via S100A1-Mediated Transient Interaction

doi: 10.1021/acschemneuro.5c00086

Figure Lengend Snippet: Structure and cross-interaction of S100A1 and S100A8 proteins. (A) CD spectra of S100A1, S100A8, and a mixture of both proteins after 70 h of aggregation at 42 °C. (B) Predicted AlphaFold model of S100A1 and S100A8 heterodimer compared to S100A8/S100A9 heterodimer (PDB ID: 1XK4 ). Time-domain DEER signals and regularized distance distributions of S100A8/S100A9 (C, D) and S100A1/S100A8 (E, F) samples at different calcium concentrations. MTSSL spin-labeled proteins are indicated by an asterisk (*).

Article Snippet: S100A1 protein is highly prevalent in the heart; however, it is also located in the brain, rivaling transcriptional levels to those of S100B, according to The Human Protein Atlas ( B), whereas S100A8 and S100A9 expression rises during neuroinflammation.

Techniques: Circular Dichroism, Labeling