Diffusion of a soluble protein, photoactivatable GFP, through a sensory cilium

Transport of proteins to and from cilia is crucial for normal cell function and survival, and interruption of transport has been implicated in degenerative and neoplastic diseases. It has been hypothesized that the ciliary axoneme and structures adjacent to and including the basal bodies of cilia impose selective barriers to the movement of proteins into and out of the cilium. To examine this hypothesis, using confocal and multiphoton microscopy we determined the mobility of the highly soluble photoactivatable green fluorescent protein (PAGFP) in the connecting cilium (CC) of live Xenopus retinal rod photoreceptors, and in the contiguous subcellular compartments bridged by the CC, the inner segment (IS) and the outer segment (OS). The estimated axial diffusion coefficients are DCC = 2.8 ± 0.3, DIS = 5.2 ± 0.6, and DOS = 0.079 ± 0.009 µm2 s−1. The results establish that the CC does not pose a major barrier to protein diffusion within the rod cell. However, the results also reveal that axial diffusion in each of the rod’s compartments is substantially retarded relative to aqueous solution: the axial diffusion of PAGFP was retarded ∼18-, 32- and 1,000-fold in the IS, CC, and OS, respectively, with ∼20-fold of the reduction in the OS attributable to tortuosity imposed by the lamellar disc membranes. Previous investigation of PAGFP diffusion in passed, spherical Chinese hamster ovary cells yielded DCHO = 20 µm2 s−1, and estimating cytoplasmic viscosity as Daq/DCHO = 4.5, the residual 3- to 10-fold reduction in PAGFP diffusion is ascribed to sub-optical resolution structures in the IS, CC, and OS compartments

Dysfunction of Heterotrimeric Kinesin-2 in Rod Photoreceptor Cells and the Role of Opsin Mislocalization in Rapid Cell Death

Loss of kinesin-2 function causes rapid death of rod photoreceptors. The cell death is dependent on the expression of opsin, which first accumulates along the route to the outer segment, but not on signaling by opsin-arrestin complexes or by light activation; the key element appears to be the accumulation of excessive protein in the wrong place

Correlations between blood and tissue omega-3 LCPUFA status following dietary ALA intervention in rats

The aim of this study was to assess relationships between the fatty acid contents of plasma and erythrocyte phospholipids and those in liver, heart, brain, kidney and quadriceps muscle in rats. To obtain a wide range of tissue omega-3 (n-3) long chain polyunsaturated fatty acids (LCPUFA) we subjected weanling rats to dietary treatment with the n-3 LCPUFA precursor, alpha linolenic acid (ALA, 18:3 n-3) for 3 weeks. With the exception of the brain, we found strong and consistent correlations between the total n-3 LCPUFA fatty acid content of both plasma and erythrocyte phospholipids with fatty acid levels in all tissues. The relationships between eicosapentaenoic acid (EPA, 20:5 n-3) and docosapentaenoic acid (DPA, 22:5 n-3) content in both blood fractions with levels in liver, kidney, heart and quadriceps muscle phospholipids were stronger than those for docosahexaenoic acid (DHA, 22:6 n-3). The strong correlations between the EPA+DHA (the Omega-3 Index), total n-3 LCPUFA and total n-3 PUFA contents in both plasma and erythrocyte phospholipids and tissues investigated in this study suggest that, under a wide range of n-3 LCPUFA values, plasma and erythrocyte n-3 fatty acid content reflect not only dietary PUFA intakes but also accumulation of endogenously synthesised n-3 LCPUFA, and thus can be used as a reliable surrogate for assessing n-3 status in key peripheral tissues.W.C. Tu, B.S. Mühlhäusler, L.N. Yelland, R.A. Gibso

Intraperitoneally injected melanin is highly uveitogenic

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