TGF β-Smad had wide signaling pathways
involved in the regulation of various life processes, such as cell growth,
differentiation, apoptosis, extracellular matrix formation and development. In
these processes, Smad4 played a core role. Research shows tumorigenesis is
associated with Smad4.
The domain (CCPmodule) of Complement control
protein (CCP) was first discovered in 1987. There are two categories of
proteins with CCP domain, regulators of complement activation (RCA) and non-regulators
of complement activation (non - RCA). The CCP domain has an important function
which can be combined with antigen molecules involved in cell signal
recognition.
CCP22 is a new and not reported gene gained
by using yeast two hybrid screening from DNA Library of human breast interacted
with Smad4. The full-length of CCP22 gene is 4494bp, encoding a protein of 1497
amino acid residues. With existing data combined bioinformatics analysis, it is
found that the protein encoded by this gene contains 21 CCP elements and 2.5
CCP sequence elements. So the gene was named CCP22.
Protein with different CCP domains, its
domains are relatively conservative. But there are some differences in the
number of them. Research shows that the proteins with CCP domains may also contain
a transmembrane domain, receptor binding domain and other domains. Unlike other
proteins with CCP domains, CCP22 only has a CCP domain. So in the CCP family of
proteins, it is not clear about its ownership. The study on CCP22 contributes
to the understanding of Smad4. It is possible to associate tumor development with
immune complement, providing new molecular targets for drug therapy. To this
end, small Interference RNA
Knockdown is constructed. Western blot shows that siRNA can effectively
reduce the expression of CCP22 protein.
At present, there are 3 main ways to get
siRNA, chemical synthesis, in vitro transcription and siRNA recombinant
expression vector construction. PSilencer2 is choosing as the siRNA expression
vector. 1－U6－neo has RNApolIII promoter, and has
the efficient, cheap and convenient advantages. The FlagCCP22 recombinant
expression vector and CCP22siRNA recombinant vector are transfected into
HEK293T cell line. Western blot shows that CCP22siRNA can effectively reduce
the protein expression level of FlagCCP22. At the same time, CCP22siRNA can
also reduce the endogenous CCP22 protein expression level, to lay a solid
foundation for further study on the function of CCP22 protein.
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