Members of the Rail, Maritime and Transport union at Southern, Merseyrail and Arriva Trains North mounted picket lines outside stations as services were disrupted or cancelled.
Drivers on Merseyrail, who are in the Aslef union, were not expected to cross picket lines, leaving managers to drive trains.
The Southern walkout was the 33rd day of action by the RMT in a row which started more than a year ago.
General secretary Mick Cash said: "Once again this morning safety-critical staff on Southern Rail are being forced by this rogue rail operator to take action in defence of safe and accessible services.
"It is a disgrace that Southern have chosen to continue their war on staff and passengers alike rather than negotiate a settlement that protects the principle of safe railways for all.
"RMT members on Southern, Northern and Merseyrail remain absolutely rock-solid and determined in separate disputes that have the common objective of putting safe rail travel before private profit.
"The total failure of these profiteering rail companies, egged on and encouraged by their political masters in the minority Tory Government, to make a single move to resolve these disputes, speaks volumes.
"RMT is available for serious talks but every approach we make is kicked back in our faces.
"It's time for the rail companies and the Government to get out of the bunker and get talks on that address the fundamental issues of safety, accessibility and the guard guarantee."
Arriva Trains North said it operated an average of more than 40% of services during the first two days of industrial action over the weekend.
Regional director Sharon Keith said: "It has been a challenging and busy weekend across the network, but by the close of business on Sunday we had been able to run more than 1,400 rail services.
"We also operated hundreds of rail replacement buses and had ticket acceptance in place on many other train operators. This all meant the north of England - and those visiting the region for special events and to enjoy the good weather - has been kept on the move.
"As we approach the Monday morning peak I would like to remind customers to continue to plan carefully. We will be operating a reduced timetable and expect all services to be extremely busy, particularly at peak times.
"The majority of services will operate between 7am and 7pm, though they will start to wind down from around 5pm."
Southern said it did not expect the RMT strike to have any effect on services, which are currently being disrupted by an overtime ban by drivers in Aslef.