Additional file 1 of A biophysical model of supercoiling dependent transcription predicts a structural aspect to gene regulation

Supporting Information. The SI of this paper is a brief derivations of the free energy of the DNA with supercoiling incorporated, an alternative derivation of the biophysical model with the supercoiling influencing the rate k kat and an application of Eq. 2 from the biophysical model in future studies. (PDF 237 kb

Cytochrome c₂ Exit Strategy: Dissociation Studies and Evolutionary Implications

Small, water-soluble, type c cytochromes form a transient network connecting major bioenergetic membrane protein complexes in both photosynthesis and respiration. In the photosynthesis cycle of Rhodobacter sphaeroides, cytochrome c2 (cyt c2) docks to the reaction center (RC), undergoes electron transfer, and exits for the cytochrome bc1 complex. Translations of cyt c2 about the RC−cyt c2 docking interface and surrounding membrane reveal possible exit pathways. A pathway at a minimal elevation allowed by the architecture of the RC is analyzed using both an all-atom steered molecular dynamics simulation of the RC−cyt c2 complex and a bioinformatic analysis of the structures and sequences of cyt c. The structure-based phylogenetic analysis allows for the identification of structural elements that have evolved to satisfy the requirements of having multiple functional partners. The patterns of evolutionary variation obtained from the phylogenetic analysis of both docking partners of cyt c2 reveal conservation of key residues involved in the interaction interfaces that would be candidates for further experimental studies. Additionally, using the molecular mechanics Poisson−Boltzmann surface area method we calculate that the binding free energy of reduced cyt c2 to the RC is nearly 6 kcal/mol more favorable than with oxidized cyt c2. The redox-dependent variations lead to changes in structural flexibility, behavior of the interfacial water molecules, and eventually changes in the binding free energy of the complex

Simulated cone mosaic produced by the quantitative model.

Simulated cone photoreceptor mosaic generated by the quantitative model displaying expression of S-opsin (blue), and M-opsin (green). A dorsal to ventral region is shown. (A1, B1, C1, D1) Complete simulated D-V strip. (A2, B2, C2, D2) Zoom in the dorsal region. (A3, B3, C3, D3) Zoom in the central region. (A3, B3, C3, D3) Zoom in the ventral region. (A1-4) S-opsin only cones. (B1-4) S-opsin expression in CEC cones. (C1-4) M-opsin expression in CEC cones. (D1-4) S- and M-opsin expression in CEC cones (E1-4) All cones including S-opsin only and CEC cones.</p

ThrB2Δ mouse Intensity Plots.

Relative intensity of S-opsin cone cells (X-axis) displayed as a function of dorsal to ventral position. Each point is colored according to the log10[Probability] of expression levels. (A) Control Retina, as seen in Fig 5F. (B) Thrβ2Δ retina.</p

Analysis of opsin expression intensity across the mouse retina.

(A, E, I) Whole mounted C57BL/6 mouse retina stained for M-opsin (green) and S-opsin (blue). (B, F, J) Heatmap displaying the log relative density of pixels that have opsin signal identified in a 25 mm2 region. (A) M-opsin signal. (B) Heatmap of total M-opsin density bins. (C) Graph of the relative density of pixels that are expressing M-opsin summed horizontally (D—V). (D) Graph of the relative density of pixels that are expressing M-opsin summed vertically (T—N). (E) S-opsin signal. (F) Heatmap of total S-opsin density bins. (G) Graph of the relative density of pixels that are expressing S-opsin summed horizontally (D—V). (H) Graph of the relative density of pixels that are expressing S-opsin summed vertically (T—N). (I) M-opsin and S-opsin (co-expression) signal. (J) Heatmap of co-expressing opsin density bins. (K) Graph of the relative density of pixels that are co-expressing S- and M-opsin summed horizontally (D—V). (L) Graph of the relative density of pixels that are co-expressing S- and M-opsin summed vertically (T—N). T = Temporal, N = Nasal, D = Dorsal, V = Ventral.</p

Modeling binary and graded cone cell fate patterning in the mouse retina - Fig 6

Model for cone cell fate specification (A) A naïve cell (grey) makes a binary decision between S-opsin only (blue) or co-expressing competent (CEC) cone fate (green, cyan or blue). The CEC cone expresses graded levels of M- and S-opsin dependendent on the D-V concentration of thyroid hormone. (B1-4) T3 (Thyroid hormone), Thrβ2* (active Thrβ2 binding T3), FD (fate determinate function), U (undifferentiated cell), S (S-only cone), C (Co-expressing cone), H (Hill function), ϕ (degradation constant of opsin proteins). (B1) Binding of T3 to Thrβ2 activates Thrβ2 (Thrβ2*) (B2) Thrβ2 controls the binary decision between S-opsin only/FD(S) or CEC/FD(C) cone fate (B3) Thrβ2* promotes M-opsin expression (B4) Thrβ2* inhibits S-opsin expression, whereas inactive Thrβ2 promotes S-opsin expression.</p

S- and M-opsin intensities in cones.

(A—B) Clustering analysis of cone populations. Cluster one = dark blue; cluster two = maroon. (C-H) Cones are ranked according to the intensity of S- and M-opsin expression levels. Intensity values are represented in arbitrary units. Each point is colored according to the log10[Probability] of expression levels. A line is drawn on the graph to show the separation between the two discrete populations of S-opsin only and CEC cone populations. (C) All cones in the regions imaged. (D) Cones in the dorsal 500–750 mm. (E) Cones in the dorsal 1500–1750 mm. (F) Cones in the central 2500–2750 mm. (G) Cones in the ventral 3500–3750 mm. (H) Cones in the ventral 4500–4750 mm.</p

Intensity of M- and S-opsins in cones.

Relative intensity of M- or S-opsin in a cone population (X-axis) is displayed as a function of dorsal to ventral position. Each point is colored according to the log10[Probability] of expression levels. (A-E) Relative intensity of M-opsin expression (F-J) Relative intensity of S-opsin expression (A, F) All M-opsin expressing cells. (B, G) All S-opsin expressing cells. For (B), arrow heads mark two distinct groups of cells in the dorsal region. (C, H) CEC cones co-expressing both S- and M-opsins. (D, I) M-opsin only expressing CEC cones. (E, J) S-opsin only expressing CEC cones.</p

Spatial distribution of M- and S-opsins in cone cells.

Relative density of a cone population summed horizontally across the image and displayed in the dorsal to ventral position. Dotted line represents midpoint of transition zone. (A) All M-opsin expressing cells. (B) All S-opsin expressing cells. (C) M-opsin only expressing cells. (D) S-opsin only expressing cells. (E) Co-expressing cells.</p

Cytochrome c₂ Exit Strategy: Dissociation Studies and Evolutionary Implications

Small, water-soluble, type c cytochromes form a transient network connecting major bioenergetic membrane protein complexes in both photosynthesis and respiration. In the photosynthesis cycle of Rhodobacter sphaeroides, cytochrome c2 (cyt c2) docks to the reaction center (RC), undergoes electron transfer, and exits for the cytochrome bc1 complex. Translations of cyt c2 about the RC−cyt c2 docking interface and surrounding membrane reveal possible exit pathways. A pathway at a minimal elevation allowed by the architecture of the RC is analyzed using both an all-atom steered molecular dynamics simulation of the RC−cyt c2 complex and a bioinformatic analysis of the structures and sequences of cyt c. The structure-based phylogenetic analysis allows for the identification of structural elements that have evolved to satisfy the requirements of having multiple functional partners. The patterns of evolutionary variation obtained from the phylogenetic analysis of both docking partners of cyt c2 reveal conservation of key residues involved in the interaction interfaces that would be candidates for further experimental studies. Additionally, using the molecular mechanics Poisson−Boltzmann surface area method we calculate that the binding free energy of reduced cyt c2 to the RC is nearly 6 kcal/mol more favorable than with oxidized cyt c2. The redox-dependent variations lead to changes in structural flexibility, behavior of the interfacial water molecules, and eventually changes in the binding free energy of the complex