"Role of the neuronal protein Cap23 in the maturation and maintenance of dendritic arbors in-vivo"

Dendrites in the central nervous system are the postsynaptic counterparts in the
neural circuitry, and the principal sites of excitatory synaptic inputs. Little is
known about the genetic elements regulating the specification, formation,
development, and maintenance of these structures. They are formed early in
development, and barring small changes in structure, remain essentially
unchanged throughout life. More than 90% of the synapses in the brain are
located on the heads and necks of dendritic protuberances called spines. Since
synapses are the functional units of brain function, a detailed study of their
anatomical and morphological features is important for understanding the
functioning of the brain, both in health and disease. But the high density and the
structural complexity of neurons, and the small size of the spines in the brain
impedes a detailed examination of spine morphology and synaptic structure.
We used a line of transgenic mice expressing membrane targeted GFP (m-GFP)
under the Thy-1 promoter to study dendritic morphology. These lines of mice
express GFP in a subset of neurons, and lights up their entire arbor, enabling
visualization of the proverbial “tree from the forest”. Expression of GFP in the
membrane gets rid of all artifacts associated with volume and intensity, and
enables visualization of fine structure of dendrites with an unprecedented clarity
and resolution. Using deconvolution confocal microscopy, we are able to detect
dendritic structures hitherto visible only in electron micrographs, and are able to
resolve spines that are below the theoretical limit of resolution of a light
microscope. This line of mice has the potential to become an invaluable assay
tool for detecting early onset defects in neuropsychiatric disorders, as it is
increasingly becoming apparent that changes in synapses (i.e. spines) are the
first markers of all neural diseases.
We next used the increased clarity offered by the m-GFP mice to address the
role of the neuronal protein Cap23 in the maintenance of dendrites. Cap23 is a
major cortical-cytoskeleton associated and calmodulin protein binding protein that
is widely and abundantly expressed during development, maintained in selected
brain regions in adults, and re-induced during nerve regeneration. Mice deficient
in Cap23 start out having normal dendritic structure and arborization, but
subsequently start decreasing in their arborization from around the time
corresponding to synapse elimination in the CNS. This decrease in branching is
progressive in nature, and correlates with the levels of the protein expressed.
Since dendrites are normal to start with, but decrease in their arborization
subsequently and in a steady manner, we refer to this novel phenomenon as
“Dendritic Atrophy”, and implicate Cap23 in the maintenance of dendrites. The
atrophy starts in the higher-order branches and proceeds towards the lower-order
ones, and the remaining branches develop ‘complex spines’.
Deficiency of Cap23 leads to the misregulation of a number of important genes in
the proteome of the brain, but not in the transcriptome, suggesting the role of
Cap23 in regulating dendritic structure by modulating the levels of several
important dendritic proteins. Interestingly, in the background of a deficiency of
Cap23, the transcriptome of the brain shows the predominant upregulation of a
number of non-coding RNAs of unknown function that show important similarities
with microRNAs. At least one miRNA (miR-128) is starkly downregulated in
Cap23 mutants. This leads to the interesting possibility that Cap23 might be
involved in the maintenance of dendrites through miRNA mediated regulation of
protein levels. Since defects in dendritic structure and arborization is a hallmark
of all neuronal diseases, it becomes interesting to speculate whether aberrations
in RNA mediated control is a general mechanism underlying neuropsychiatric
diseases in general.