Category Archives: 14CoS ko, hepatocytes, rescue with NTBC

14CoS/14CoS mice homozygous for a 3800 kb deletion on chromosome 7, electron microscopy NTBC rescue

Nucleolar ultrastructure: fibrillar center organization

May 29, 201714CoS ko, hepatocytes, rescue with NTBC, Electron micrographs of liver, Nucleoar organization, The cell: images, Ultimate order, the cellthankusc

I was looking at this micrograph of a n hepatocyte with the fibrillar centers and dense thick fibrils in their midst, and the granular zone around the outside of this nucleolus. The latter being very punctate in appearance, just really obvious. I measured these once again at about 23 nm in diameter and noticed that around many of the fibrilar centers where was a basket, or ring, or radial distribution of these small punctate appearances. OK so they are not really punctate, but can appear smeared like fibrils too, though the granular appearance and the nomenclature is well entrenched in the literature, they are just going to be strands not dots. It could be that these radial dots around the fibrillar centers are bands, or windings at a given distance apart, kind of like fingers of a basket, or more aptly put, like a cage, with wires of 23 nm in diameter.

Electron micrograph here shows a portion of a nucleolus with three fibrillar centers cut out and enlarged (original sites are boxed in white), and a portion of the outer nuclear membrane with ribosomes (for measurement – one ribosome being about 27 nm in diameter, top right image with red dot and text). The cross section of the cage, or basket filaments around the fibrillar centers are shown as black dots.

Estimates of the size of some nucleolar components

May 26, 201714CoS ko, hepatocytes, rescue with NTBC, Electron micrographs of liver, Nuclear pores and nuclear architecture, Nucleoar organization, Ultimate order, the cellthankusc

This micrograph, parts of which have been seen before, is an hepatocyte nucleus which is in the early stages of demise (untreated CoS14 ko without rescue). The nucleolus is large, and has two cajal type bodies (one seen here) and fibrillar centers with fairly large densities within (of approximately 90 nm diameter). I have given these sizes (relative to a ribosome size red dot approximately 27 nm (lower right hand side of micrograph labeled as such). The granules within the granular portion of the nucleolus (as well as the banding patterns found on cajal bodies) is something around 23 nm. What I presume to be perichromatin granules look to be about 50 nm. The large dense bodies within the fibrilar centers are a little unusual.

Nuclear pores: diagrams vs micrographs

May 15, 201714CoS ko, hepatocytes, rescue with NTBC, Nuclear pores and nuclear architecture, The cell: images, Ultimate order, the cellthankusc

It is sometimes difficult to reconcile all the graphics that appear in research articles with what actually one sees with the electron microscope. There are artists and scientists trying to communicate facts and abstract ideas and the results are something hysterical.

Anyway, in looking at the 14CoS ko 48 hr no rescue nucleus in an hepatocyte I found this nuclear pore which actually had what looks like some of the 8-mer basket proteins that were quite clear. I have highlighted those (on the nuclear side of the pore) in blue and highlighted the cytoplasmic side with cytoplasmic filaments in rust color. One can see from a diagram from wikipedia (thank you wikipedia) which I have extensively simplified and modified, that the original and what I have seen in actuality don’t really fit. I cut the diagram apart and extended the inner nuclear membrane space, shrunk up the basket and extended the cytoplasmic filaments (see diagram on the right). The other issue is likely that the micrograph tends to show the pore from the ” cytoplasmic viewpoint” or bottoms up, while the diagram looks more “at the horizontal level”. Basically, the nuclear pore complex is an absolute wizard structure… fun to learn about. This electron micrograph is from a 48 hr old 14CoS ko pup, not rescued with NTBC. 18750_80206_anim#35 liver. More posts on this topic HERE.

What i have trouble visualizing with electron microscopy are the three rings…. the cytoplasmic ring, the nuclear side ring and that ring structure that individuals claim is at the rounded fusion of the trilaminar inner and outer nuclear membranes.

Here is another nuclear pore from the same electron micrograph in which I have highlighted the four areas which perhaps will correspond to the nuclear side ring 2 dots at the bottom (along with light blue arrow of some particle just on the way out of the nucleus likely) and the cytoplasmic side ring above). Orange arrow points to where the trans membrane ring should be seen.

Fibril, filament, snRNPs, granular component etc

May 9, 201714CoS ko, hepatocytes, rescue with NTBC, Electron micrographs of liver, Nuclear pores and nuclear architecture, Nuclear symmetry during apoptosis, Nucleoar organization, The cell: images, Ultimate order, the cellthankusc

This micrograph I am using to determine the sizes of the different nuclear structures, so this is the first attempt to define at least four different sizes of components. For me, there is similarity in the size of the granules (beads on a string maybe something like 30 nm) in the cajal body and in the nucleolus. Arrows point to the kind of layered banding parallel strand look of the cajal bodies, the red circle is about the size of a cytoplasmic ribosome (which would be something like 27nm) and the diameter of the rounded object beside the cajal body (something on the order of 60nm) and an even larger fibrillar component seen through out the fibrillar centers and also just a little above center left (orange spot) which might be 130 – 150nm. So there are four measurements of fibrillar components in this nucleus, so far. Will post more.

Nuclear pores: more than usual

May 4, 201714CoS ko, hepatocytes, rescue with NTBC, Electron micrographs of liver, Nuclear pores and nuclear architecture, The cell: images, Ultimate order, the cellthankusc

Hepatocyte from a 14CoS -/- mouse (homozygous for a 3800kb deletion on chromosome)that has spent 360 days being rescued from certain demise by NTBC has little condensed chromatin, very round and active looking nucleus and literally thousands of nuclear pores, more than I remember seeing in the nuclear envelope of any other cell type, and any other experimental condition. Totally awesome. I would not hazard a guess as to how many are there per micron sq.

The purpose of nuclear pores is of course do provide a mechanism for selective transport of molecules in and out of the nucleus, separating the compartment of the cytoplasm and nucleus. All produced proteins from the cytoplasm required by the nucleus will be imported through the nuclear pore, and the RNAs, mRNA, tRNA and rRNA, destined to become cytoplasmic machinery, transcribed and modified (post-transcriptional modification) in the nucleus will be exported to the cytoplasm through nuclear pores.

So the electron micrograph below (and inset) show this massive number of nuclear pores, particularly on one side that is cutting through the nuclear membrane at a slightly tangential angle, and also peculiarly, the polysomes (clear spirals of mRNA and rRNA) on the cytoplasmic side of the outer nuclear membrane are just about the only place in the cytoplasm where one finds RER. This vesicular ultrastructure (and particularly the vesicle – within – a vesicle) has been published Deiter et al 14cos, but not the nuclear pore observation. Here is such an hepatocyte nucleus, and inset to show tangentially cut nuclear pores (arrows), and spirals of mRNA with ribosomes (arrowheads). 17709_65053_14CoS-/- 360d w NTBC.

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