Du Plessis finished unbeaten on 78 and guided crucial partnerships of 57 with Robin Peterson (31) and 64 with Vernon Philander (28) to lift South Africa out of serious trouble at 75-6 after winning the toss.
Australia reached 33-2 in the 11 overs before stumps, with David Warner unbeaten on 12 and nightwatchman Nathan Lyon 5 not out.
Ricky Ponting, playing his 168th and last Test match, waited in the pavilion for 22 minutes after Lyon went in at the fall of Watson’s wicket.
The Australian reply started badly when Dale Steyn removed Ed Cowan (0) on the third ball of the innings and Philander picked up his first wicket in the series when he got an lbw decision against Shane Watson (10) on a referral, overturning umpire Asad Rauf’s initial not out call, as the hosts slumped to 18-2.
A dozen wickets fell in two clusters on a fast, bouncy WACA pitch when conditions were overcast in the morning and evening sessions.
Lyon was the unexpected contributor for Australia, with his offspin delivering 3-41 for a new-look bowling attack missing all three pacemen who played in the first two drawn Tests.
Mitch Starc, who was 12th man in Brisbane and Adelaide, took two big wickets immediately before lunch and another left-armer, Mitchell Johnson, returned 2-54 in his first Test in a year.
The pacemen were on top early and South Africa was in a desperate position after losing five wickets for 14 runs before allrounder Peterson and then Philander helped du Plessis resurrect the innings.
Philander batted aggressively, hitting a big six off Lyon and two boundaries before he skied the Australia offspinner to Mike Hussey, who took a well-judged catch at deep mid-on. Steyn (2) chopped a delivery from Johnson onto his stumps to make it 206-9.
Morne Morkel (17) stroked four boundaries off Johnson before he miscued Lyon to John Hastings at mid-on to end the innings.
Du Plessis was again the big obstacle for the Australian bowlers four days after he batted through the final day and finished unbeaten on 110 to save the second Test on debut at Adelaide on Monday. He has 266 runs in three innings, and 188 since his last dismissal, after earning his call up when JP Duminy was injured in a training accident after the opening day of the first Test at Brisbane.
He batted for seven hours in his second innings in Adelaide and was unbeaten again in 3 hours, 15 minutes on Friday, facing 142 balls and hitting 12 boundaries after going to the crease with his team reeling at 67-5 in the 28th over.
The South Africans started the Test positively after winning the toss and had only lost skipper Graeme Smith (16) as they progressed to 61-1 before Starc struck twice to make it 63-3 at lunch when he bowled opener Alviro Peterson (30) and Jacques Kallis (2) with swinging deliveries.
Hashim Amla (11) had a reprieve five balls after lunch when Cowan put down a regulation chance off Hastings’ bowling. But he was out next over, without adding to his score, when he was run out at the striker’s end by David Warner’s direct hit.
Hastings got a ball to swing out and late in his next over and picked up his first Test wicket when AB de Villiers (4) edged to Clarke at slip and South Africa slumped to 67-5.
Dean Elgar faced 12 balls on debut before attempting to pull a sharply rising ball from Johnson and top-edging down leg side to wicketkeeper Matthew Wade, and Lyon ended the seventh-wicket stand when he had Peterson caught behind while attempting a cut shot.
South Africa needs only to draw the Perth Test to retain the No. 1 Test ranking. Australia needs a win to replace the tourists at No. 1 and has extra motivation for winning after Ponting announced this match would be his last.
Brief Scores: South Africa 225 all out (Faf du Plessis 78 n.o, R Petersen 31, N Lyon 3/41, M Johnson 2/51) vs Australia 33/2 (D Warner batting 12, V Philander 1/8)