Temporal Dysynchrony in brain connectivity gene expression following hypoxia

List of K-means cluster analysis of connectivity genes across development during hypoxia. Relative log2 fold change compared to the developmental average is provided. (XLSX 197 kb

Effect of conditional deletion of cytoplasmic dynein heavy chain DYNC1H1 on postnatal photoreceptors.

Cytoplasmic dynein (dynein 1), a major retrograde motor of eukaryotic cells, is a 1.4 MDa protein complex consisting of a pair of heavy chains (DYNC1H1) and a set of heterodimeric noncatalytic accessory components termed intermediate, light intermediate and light chains. DYNC1H1 (4644 amino acids) is the dynein backbone encoded by a gene consisting of 77 exons. We generated a floxed Dync1h1 allele that excises exons 24 and 25 and truncates DYNC1H1 during Six3Cre-induced homologous recombination. Truncation results in loss of the motor and microtubule-binding domain. Dync1h1F/F;Six3Cre photoreceptors degenerated rapidly within two postnatal weeks. In the postnatal day 6 (P6) Dync1h1F/F;Six3Cre central retina, outer and inner nuclear layers were severely disorganized and lacked a recognizable outer plexiform layer (OPL). Although the gene was effectively silenced by P6, DYNC1H1 remnants persisted and aggregated together with rhodopsin, PDE6 and centrin-2-positive centrosomes in the outer nuclear layer. As photoreceptor degeneration is delayed in the Dync1h1F/F;Six3Cre retina periphery, retinal lamination and outer segment elongation are in part preserved. DYNC1H1 strongly persisted in the inner plexiform layer (IPL) beyond P16 suggesting lack of clearance of the DYNC1H1 polypeptide. This persistence of DYNC1H1 allows horizontal, rod bipolar, amacrine and ganglion cells to survive past P12. The results show that cytoplasmic dynein is essential for retina lamination, nuclear positioning, vesicular trafficking of photoreceptor membrane proteins and inner/outer segment elaboration

Arf-like Protein 2 (ARL2) Controls Microtubule Neogenesis during Early Postnatal Photoreceptor Development

Arf-like protein 2 (ARL2) is a ubiquitously expressed small GTPase with multiple functions. In a cell culture, ARL2 participates with tubulin cofactor D (TBCD) in the neogenesis of tubulin αβ-heterodimers, the building blocks of microtubules. To evaluate this function in the retina, we conditionally deleted ARL2 in mouse retina at two distinct stages, either during the embryonic development (retArl2−/−) or after ciliogenesis specifically in rods (rodArl2−/−). retArl2−/− retina sections displayed distorted nuclear layers and a disrupted microtubule cytoskeleton (MTC) as early as postnatal day 6 (P6). Rod and cone outer segments (OS) did not form. By contrast, the rod ARL2 knockouts were stable at postnatal day 35 and revealed normal ERG responses. Cytoplasmic dynein is reduced in retArl2−/− inner segments (IS), suggesting that dynein may be unstable in the absence of a normal MTC. We investigated the microtubular stability in the absence of either ARL2 (retARL2−/−) or DYNC1H1 (retDync1h1−/−), the dynein heavy chain, and found that both the retArl2−/− and retDync1h1−/− retinas exhibited reduced microtubules and nuclear layer distortion. The results suggest that ARL2 and dynein depend on each other to generate a functional MTC during the early photoreceptor development

Arf-like Protein 2 (ARL2) Controls Microtubule Neogenesis during Early Postnatal Photoreceptor Development

Arf-like protein 2 (ARL2) is a ubiquitously expressed small GTPase with multiple functions. In a cell culture, ARL2 participates with tubulin cofactor D (TBCD) in the neogenesis of tubulin αβ-heterodimers, the building blocks of microtubules. To evaluate this function in the retina, we conditionally deleted ARL2 in mouse retina at two distinct stages, either during the embryonic development (retArl2−/−) or after ciliogenesis specifically in rods (rodArl2−/−). retArl2−/− retina sections displayed distorted nuclear layers and a disrupted microtubule cytoskeleton (MTC) as early as postnatal day 6 (P6). Rod and cone outer segments (OS) did not form. By contrast, the rod ARL2 knockouts were stable at postnatal day 35 and revealed normal ERG responses. Cytoplasmic dynein is reduced in retArl2−/− inner segments (IS), suggesting that dynein may be unstable in the absence of a normal MTC. We investigated the microtubular stability in the absence of either ARL2 (retARL2−/−) or DYNC1H1 (retDync1h1−/−), the dynein heavy chain, and found that both the retArl2−/− and retDync1h1−/− retinas exhibited reduced microtubules and nuclear layer distortion. The results suggest that ARL2 and dynein depend on each other to generate a functional MTC during the early photoreceptor development

Additional file 10: Figure S2. of Temporal Dysynchrony in brain connectivity gene expression following hypoxia

Larger file and font sizes showing gene names for the Protein-Protein Interactions Network. A) STRING analysis of most significant (adjusted p < 0.05) genes interactions, n = 57; color key for interaction type is shown to the right. B) STRING analysis with relaxed criteria (unadjusted p < 0.05), n = 244. (JPG 2997 kb

Additional file 2: of Temporal Dysynchrony in brain connectivity gene expression following hypoxia

List of K-means cluster analysis of connectivity genes across development during normoxia. Relative log2 fold change compared to the developmental average is provided. (XLSX 171 kb