Page 1 \ ?^p 6r.1 CELL CYCLE CONTROL OF HUMAN H4 ...

A1i2 F: Ceu a:rcte cotuot ot H1 G@ ftqNtioti@
transcrip on lactors binding ro mutant prcmot€rs do not hav€ dominant
efiects on dete.mininq tlre level of reports. geno transcnplion- This tunclional
flenbilty in nucleotde spacing is consst6nt with sub!€ del€tions and
insertio.s niat appesr to have occud€d duing €volulion ol th€ cons€v€d
sequences in sle ll(vanwinen ota/., 1992).
Ta.gsled murarlon olrhe HINF-M/|nF.2 blnding site reduces H4
hlroduction of rwo substiturion mutarions {mutanl MsP-16) precisely
designed in lhe rsgion ol th. M.box lhat doss nol ovenap wilh lhe H|NF.
P/H4IF 2 binding siie, pre!€nts binding ol HiNF-M/|RF-2 bur nor HiNF-
P/HaTF-2 o. H|NF O b Sne ll based on g€r shitr analysis (Fig. 27). This
mltation and additional substitullon ol three nucleotides in lhe NGGTCCGNN
molif {mutant MPIM-17) abolishes int€raclions of both H|NF lMllFF 2 and
HiNF-P/H4-TF2 wir.,i Sile ll, whil€ reducifg the HiNF-D/Site Ll inleraction (Fig.
27). Slbslitution ot rwo guanine rssidles (mutant lGlM-18) representing
methylation intederence .ontacts lo. both HiNF-M/|FF.2 and HINF-P/H4-TF2
abolishes HiNF-P/H4-TF2 and rgducss binding of HiNF-rM/lRF-2 and HiNF-O
ro si16 [ (Fig.27).
Comparison oJ reponer gen€ erprossion with lhese construds relative
to the wildlype H4 promoter reveals lhar th€ HiNF M/IRF 2 mdanl MSPno
conslruct displays 2 10 3 lold reducsd promoter act'vity (Flg. 30). Ihe
quanlitative resembla.ca of rhs mutarlonal €lrsct wirh ihe 3 fold reducrion
obserued wllh rhe olher rwo HiNF-M/IFF-2 mutanl conslructs iNs-lo/cAT
130