Lec 2

Vocabulary practice flashcards covering plasmid partitioning mechanisms (Type I ParA/ParB and Type II ParM/ParR), plasmid addiction and toxin-antitoxin systems (including hok/sok), and the causes and outcomes of plasmid incompatibility.

Plasmid Partitioning

An active segregation process that ensures plasmids are equally distributed to opposite daughter cells prior to cell division using mitotic-like systems such as ParA/ParB or ParM/ParR.

Type I Plasmid Partitioning

A plasmid segregation system consisting of the centromere-like sequence $parS$, the DNA-binding adapter protein ParB, and the filament-forming ATPase ParA.

$$parS$$

A centromere-like cis-acting DNA site on a plasmid that serves as the specific segregation locus bound by ParB during Type I partitioning.

ParA

A filament-forming ATPase that binds the host nucleoid in its ATP-bound state; its local hydrolysis by ParB directs plasmid translocation toward areas of higher ParA concentration.

ParB

A sequence-specific DNA-binding protein that recognizes the $parS$ site and stimulates localized ATP hydrolysis of ParA-ATP to drive plasmid segregation.

Plasmid Surfing

The mechanism in Type I partitioning where interaction of ParB with ParA-ATP causes local hydrolysis into ParA-ADP, creating a depleted clearing zone behind the plasmid and pulling it toward regions of higher ParA concentration.

Type II Plasmid Partitioning

A segregation mechanism that utilizes an actin-like motor protein (ParM) and a DNA-binding adapter (ParR) to actively push paired plasmids to opposite poles of the bacterial cell.

ParM

An actin-like protein ($m = \text{motor}$) that polymerizes into a dynamic, mitotic spindle-like filament between plasmids to push them to opposite cellular poles.

ParR

A plasmid DNA-binding protein in Type II partitioning that binds to the $parC$ site and serves as the adapter that connects the plasmid to the growing ParM filament.

• like ParB

$$parC$$

The centromere-equivalent cis-acting DNA sequence on a plasmid that is specifically recognized and bound by ParR in Type II partitioning systems.

• like parS

Plasmid Addiction System

A post-segregational killing mechanism consisting of a stable toxin and an unstable antitoxin that selectively kills daughter cells that fail to inherit the plasmid

• unstable antitoxin = degrades quickly ; therfore stable toxin kills plasmid.

Toxin

A stable plasmid-encoded molecule that persists in the cell and kills it if the short-lived neutralizing antitoxin is no longer synthesized following plasmid loss.

Antitoxin

An unstable plasmid-encoded RNA or protein molecule that continually neutralizes its cognate stable toxin to ensure cell survival as long as the plasmid is retained.

Type I Toxin-Antitoxin System

A toxin-antitoxin arrangement where an unstable antisense RNA antitoxin base-pairs with a stable toxin mRNA, repressing its translation and promoting degradation by RNase III.

Hok

The Host Killing toxin encoded as a stable mRNA; once translated, the small toxin inserts into the bacterial inner membrane and disrupts membrane potential, causing cell death.

Sok

The Suppression of Killing antitoxin consisting of an unstable antisense RNA that base-pairs with $hok$ mRNA to form a double-stranded RNA substrate targeted for degradation by RNase III.

Plasmid Incompatibility

The inability of two different plasmids to be stably maintained together within the same bacterial cell lineage because they share identical replication or partitioning systems.

• recall more plasmids = more survival for bacteria

2 reasons for plasmid incompatability

1. Replication incompatability —> share same oriV

2. Partitioning incompatability —> share same Par

machinery cant differnetiate between the plasmids

Replication Incompatibility

A form of incompatibility occurring when plasmids possess identical $oriV$ sequences and regulatory components (e.g., RepA), preventing the cell from distinguishing them during copy-number regulation.

Partitioning Incompatibility

Incompatibility that arises when two plasmids share the same Par machinery, causing the partitioning apparatus to segregate them indiscriminately and leading to the loss of one plasmid line.

Plasmid Curing

The loss of a plasmid from a bacterial cell lineage, which occurs spontaneously over successive divisions or can be encouraged experimentally by omitting selective pressure.

eg. “cured” of the red plasmid = there is no more red plasmid

how could you prevent curing of a plasmid?

• increase copy number

• each have a distinct toxin/antitoxin system

• different antibiotic resistance gene + antibiotic selection

how could you encourage curing of a plasmid?

• select for one and no selection for the other