Fluorescent proteins that target PD show a punctate distribution of fluorescence along the wall of plant cells that reflect the distribution of individual PD channels or, more commonly, the distribution of groups of PD in pit fields.
Using aniline blue to stain callose [ 27], ATM1 was shown to accumulate in plasmodesmata-enriched pit fields.
When transiently expressed in N. benthamiana, GFP-ATM1(IQ-tail) was found in pit fields accumulating callose, and co-localized with the ER.
The dotted pattern seen in some of the root cells was identified by an anti myosin VIII antibody in maize roots[ 11, 15] as plasmodesmata enriched pit fields.
Higher magnification of these regions of the inner face of cell walls with Calcofluor White fluorescence (Figs. 9c-e) revealed regions with reduced fluorescence, that are possibly pit fields.
Further this work provide evidence using microscopy of live cells, that Myosin VIII is preferentially involved in the function of ARA6 associated endosomes and is localized to ER and pit fields rich in plasmodesmata.
No labelling was found throughout the cell walls and very little was detected in the middle lamella, confirming that calcium cross-linked HG epitopes are restricted to the middle lamellae at cell corners and pit fields [ 29].
ATM1 has also been found in pit fields in the inner cortex cells of maize root apices, where it was predicted to be involved in fluid-phase endocytosis [ 12].
Using a specific stain for callose we confirmed the presence of ATM1 in pit fields of N. benthamiana leaves, however, in the transgenic plants, ATM1 fluorescent dots are scattered all over the cell.
While raw data of co-staining with the callose-specific dye Anilin Blue suggests incongruent distribution domains of cellulose in callose in the cell wall, this becomes much clearer on deconvolved images, even showing the thin cell wall plate that remains in the pit fields.
