Organelle Markers: Antibodies to the Nucleus - FocusOn 098

In this edition of our series to organelle markers we get to the core. We focus on markers which are useful to stain proteins of the nucleus. Therefore, plenty of different proteins can be used from the nuclear lamina to transcription factors, histones and proteins of the nucleolus.

PARP

Poly (ADP-ribose) polymerases are nuclear matrix enzymes involved in the repair of DNA strand breaks. Their function is based on specific interactions with DNA and nuclear proteins. These enzymes modify nuclear proteins by poly-ADP-ribosylation, which is required for DNA repair, regulation of apoptosis, and maintenance of genomic stability. At the moment 17 members of the PARP family are known. Acris Antibodies provides antibodies to PARP 1, 3, 6, 11, 12 and 16 at this time.

Protein P84 / THOC1

Another excellent specific marker of the nuclear matrix is Protein P84. Using antibodies to this 84 kDa nuclear matrix protein (THOC1) is helpful in cell fractionation studies, for example. The THO/TREX complex is recruited to transcribed genes and travels with the RNA polymerase
during elongation. It links proteins that function in transcription and in RNA export.

Fig. 1: Formalin-fixed, paraffin-embedded (FFPE) human tonsil stained with PARP1 antibody Cat.-No. AP15715PU-N

Fig. 2: Human colon (FFPE) stained with THOC1 antibody Cat.-No. AM06060PU-N

Nucleoporins (NUP)

Another possibility to mark the nucleus is using antibodies to nucleoporins (NUP). The nuclear pore complex is a massive structure that extends across the nuclear envelope, forming a gateway that regulates the flow of macromolecules between nucleus and cytoplasm. Nucleoporins are the main components of the nuclear pore complex in eukaryotic cells and several different members are known like NUP50.

Fig. 3: Western blot analysis of NUP35 in human liver lysate in the 1) absence and 2) presence of immunizing peptide, 3) mouse brain lysate and 4) rat brain lysate using Cat.-No. AP22066PU-N

Fig. 4: Confocal microscope analysis of NUP153, lamin B3 and F/GXFG nucleoporin distribution in sperm pronuclei using NUP153 antibody Cat.-No. BP4506

Histones

The histone proteins serve as a structural scaffold for the organisation of nuclear DNA into chromatin. In eucaryotic cells there are 4 core histone proteins, H2A, H2B, H3 and H4, that assemble to form an octamer round which the DNA is wrapped in a sequence dependent manner. The resulting complex is known as a nucleosome and is the basic subunit of chromatin. Histone H1 binds to each nucleosome and links the nucleosomes together resulting in the 30 nm chromatin fiber and so into higher order structures.

NF-kappa-B

This pleiotropic transcription factor is present in almost all cell types. It is involved in different biological processes like inflammation, immunity, differentiation, cell growth, tumorigenesis and apoptosis. It was originally identified as a heterodimeric DNA binding protein complex consisting of p65 (RelA) and p50 (NFKB1) subunits. Other identified subunits include p52 (NFKB2), c-Rel, and RelB.

Fibrillarin

A good choice to stain the nucleolus is fibrillarin (FBL) also known as 2‘-O-ribose RNA methyltransferase.

Fig. 5: Human SH-SY5Y cells stained with fibrillarin antibody Cat.-No. AM08249PU-N, showing prominent specular nucleolar staining (nuclei counter stained with DAPI DNA stain; very pale blue)

Other markers

Further nuclear marker antibodies are available like antibodies to the transcription factor AP1/c-jun or NFKB.
Antibodies to lamin proteins (intermediate filament-type proteins which form major components of the nuclear lamina) and lamin associate proteins are available and described in FocusOn 028.
Estrogen receptors which serve as nuclear hormone receptors are described more detailed in FocusOn 050.

Further readings

See the PDF-version: FocusOn 098